Pharmaceutical compositions comprising cross-linked small molecule amine polymers

ABSTRACT

Anion-binding polymers are described. The anion-binding polymers in some cases are low swelling anion-binding polymers. In some cases, the anion-binding polymers have a pore volume distribution such that a fraction of the polymer is not available for non-interacting solutes above a certain percentage of the MW of the target ion for the polymer. In some cases, the anion-binding polymers are characterized by low ion-binding interference, where the interference is measured in, for example, a gastrointestinal simulant, relative to non-interfering buffer. Pharmaceutical composition, methods of use, and kits are also described.

CROSS-REFERENCE

This application is a continuation-in-part application of Ser. No.10/965,044 entitled ANION-BINDING POLYMERS AND USES THEREOF, filed Oct.13, 2004, which is a continuation-in-part application of Ser. No.10/806,495 entitled CROSSJANKED AMINE POLYMERS, filed Mar. 22, 2004;Ser. No. 10/701,385 entitled POLYAMINE POLYMERS, filed Nov. 3, 2003; allof which are incorporated herein by reference in their entirety and towhich applications we claim priority under 35 USC §120.

BACKGROUND OF THE INVENTION

Ion selective sorbents have been used in human therapy to correctdisorders in electrolyte balance, in conditions such ashyperphosphatemia, hyperoxaluria, hypercalcemia, and hyperkalemia.Hyperphosphatemia occurs in patients with renal failure, whose kidneysno longer excrete enough phosphate ions to compensate exogenousphosphate uptake in the diet. This condition leads to high serumphosphate concentration and high calcium x phosphate product. Althoughthe etiology is not fully demonstrated, high calcium x phosphate producthas been held responsible for soft tissue calcification andcardiovascular disease. Cardiovascular disease is the cause of death inalmost half of all dialysis patients.

Aluminum, calcium, and, more recently, lanthanum salts have beenprescribed to control phosphate ion absorption in the gastrointestinal(GI) tract and restore systemic phosphate levels back to normal. Howeverthese salts liberate soluble aluminum and calcium cations in the GItract, which are then partially absorbed into the blood stream. Aluminumabsorption can cause serious side effects such as aluminum bone diseaseand dementia; high calcium uptake leads to hypercalcemia and putspatients at risk for coronary calcification.

Metal-free phosphate binders such as strong base ion-exchangermaterials, Dowex and Cholestyramine resins, have been suggested for useas phosphate binders. However, their low capacity of binding requireshigh dosage that is not well tolerated by patients.

Amine functional polymers have been described as phosphate or oxalatebinders. For example, see 5,985,938; 5,980,881; 6,180,094; 6,423,754;and PCT publication WO 95/05184. Renagel, a crosslinked polyallylamineresin, is a phosphate sequestering material introduced in the market asa metal-free phosphate binder. In vitro phosphate binding of Renagel isapproximately 6 mmol/gm in water and 2.5 mmol/gm when measured in 100 mMsodium chloride and 20 mM phosphate at neutral pH. The recommendeddosage for the targeted patient population is typically between 5gms/day to 15 gms/day to keep the phosphate concentration below 6 mg/dLPublished phase I clinical trials on Renagel, performed on healthyvolunteers, indicate that 15 gins of Renagel decrease the phosphateurinary excretion from a baseline of 25 mmole to 17 mmole, thedifference being excreted in the feces as flee and polymer-boundphosphate From these data, the in vivo capacity range can be establishedat 0 5-1 mmol/gm, which is much less than the in vitro capacity of 6mmol/gr measured in saline Considering only the in vitro bindingcapacity of Renagel measured in saline, a dosage of 15 gm of phosphatebinder would bind more than the entire phosphorus content of the averageAmerican diet, i.e 37 mmol/day. The discrepancy between the in vitrobinding capacity and the documented low in vivo binding capacity has anegative impact on the therapeutic benefit of the drug since more resinis needed to bring the serum phosphate to a safe range.

This loss of capacity of ion-exchange resins is not limited to Renagelwhen used in the complex environment of the GI tract environment.Although generally safe from a toxicological perspective, the large doseand inconvenience associated with taking multigram amounts of resinargues for the need to improve resin capacity. As an example, even inreported safety studies of the Renagel binder, patients have notedgastrointestinal discomfort at doses as low as 1.2-2.0 gm/day for an 8week treatment period. Patients receiving 5.4 gm of Renagel/day werediscontinued from treatment due to adverse events such as GI discomfortin 8.9% of the cases (Slatapolsky, et al Kidney Int. 55:299-307, 1999;Chertow, et al Nephrol Dial Transplant 14:2907-2914, 1999). Thus, animprovement in in vivo binding capacity that translates to lower, bettertolerated dosing would be a welcome improvement in resin-basedtherapies.

As a result of these considerations there is still a great need forsafe, high-capacity binders that selectively remove ions from the bodywith a lower drug dosage and a better patient compliance profile.

Patient compliance is recognized today as one of the main limitingfactors for patients to comply with the K/DOQI recommendations: doseescalation implies that patients have to take ten 800 mg pills per dayand beyond. Renagel pills take the form of swallowable tablets and areadministered with a minimum of fluid, adding to the burden of ESRDpatients who are under fluid restriction. More easy-to-takepharmaceutical formulation would be desirable: in particular chewabletablets are becoming more popular amongst the geriatric and pediatricpopulation and in treatments requiring a large pill burden: chewabletablets allows greater strength pills and ultimately reduces the numberof tablets per meal. Because the active contained in a chewable tabletis first dispersed under the effects of mastication and saliva beforebeing swallowed, the requirements on both the shape and weight of thetablet are much less severe than those imposed on swallowable tablets:However, until now it was not possible to formulate a hydrogel such asRenagel in a chewable tablet because of the high swellingcharacteristics of that polymer: Renagel usually swells very rapidly upto about 10 times its weight in an isotonic solution. This has two muchundesired consequences: firstly, while in the mouth the polymer willswell and give a very unpleasant feel (dry mouth, sensation of choking);secondly, even if patient overcomes the sensory in mouth, theadministration of a swollen gel in the esophagus can be hazardous.Besides, it is also well known that highly swellable gels, whenadministered in the multi grams range, provoke side effects such asbloating, constipation or diarrhea.

SUMMARY OF THE INVENTION

In one aspect, the invention provides anion-binding polymers. In someembodiments, the invention provides an anion-binding polymer where thepolymer binds a target anion (e.g., phosphate or oxalate), and where thepolymer is characterized by at least two of the following features: a) aswelling ratio of less than about 5; b) a gel pore volume distributionmeasured in a physiological medium characterized by a fraction of saidpore volume accessible to non-interacting solutes, of molecular weightgreater than about twice the MW of the target anion, of less than about20% of the weight of the gel; and c) an ion-binding interference for thetarget anion lower than about 60% when measured in a gastrointestinalsimulant, relative to a non-interfering buffer. In some embodiments, theswelling ratio is less than about 4, or less than about 3, or less thanabout 2.8, or less than about 2.7, or less than about 2.6, or or lessthan about 2.5. In some embodiments, the polymer binds bile acids orcitrate with a capacity of less than about 2 mmol/gm, or less than about1 mmol/gm, or less than about 0.5 mmol/gm, or less than about 0.3 mm/gm,or less than about 0.1 mm/gm. In some embodiments, the swelling ratio ismeasured in isotonic solution and neutral pH. In some embodiments, thepolymer comprises amine monomers. In some embodiments, the aminemonomers are selected from the group consisting of allylamine,vinylamine, ethyleneimine, 1,3 diamino propane, and N,N,N′,N′-tetrakis(3-aminopropyl) 1,4 diaminobutane, 1,2,3,4 tetraaminobutane, Formula 1and Formula 2, where Formula 1 and Formula 2 are the followingstructures:

In some embodiments, the invention provides an anion-binding polymercontaining crosslinked polyamines, where the polymer is obtained byinverse suspension, and wherein the swelling ratio of the polymer isless than 5.

In some embodiments, the invention provides a phosphate-binding polymerwhere the polymer is characterized by at least one of the followingfeatures: a) a swelling ratio of less than about 5, preferably less thanabout 2.5; b) a gel pore volume distribution measured in a physiologicalmedium characterized by a fraction of said pore volume accessible tonon-interacting solutes, of molecular weight greater than about 200, ofless than about 20% of the weight of the gel; and c) an ion-bindinginterference for phosphate lower than about 60% when measured in agastrointestinal simulant, relative to a non-interfering buffer. In someembodiments, the swelling ratio is less than about 2.8, or less thanabout 2.7, or less than about 2.6. In some embodiments, the polymerbinds bile acids or citrate with a capacity of less than about 2mmol/gm, or less than about 1 mmol/gm, or less than about 0.5 mmol/gm,or less than about 0.3 mm/gm, or less than about 0.1 mm/gm. In someembodiments, the swelling ratio is measured in isotonic solution andneutral pH.

In some embodiments, the invention provides a phosphate-binding polymerwhere the polymer is characterized by a swelling ratio of less thanabout 5, preferably less than about 2.8, or less than about 2.7, or lessthan about 2.6, most preferably less than about 2.5, where this ratio ismeasured in isotonic solution and neutral pH. In embodiments, thepolymer has a mean in vivo phosphate binding capacity of greater thanabout 0.5 mole/gm. In embodiments, the polymer is a polyamine polymer,and the chloride content of the polymer is less than about 35 mol % ofthe content of amine groups.

In some embodiments, the invention provides an anion-binding polymerwhere the polymer binds a target anion (e.g., phosphate or oxalate), andwhere the polymer is characterized by at least two of the followingfeatures: a) a swelling ratio of less than about 5; b) a gel pore volumedistribution measured in a physiological medium characterized by afraction of said pore volume accessible to non-interacting solutes, ofmolecular weight greater than about twice the MW of the target anion, ofless than about 20% of the weight of the gel; and c) an ion-bindinginterference for the target anion lower than about 60% when measured ina gastrointestinal simulant, relative to a non-interfering buffer, wherethe polymer contains one or more amine monomers and one or morecrosslinkers, and where the polymer is produced by a process in whichthe amine is present in solvent before crosslinking at a ratio ofamine:solvent of from about 3:1 to about 1:3 and the total contentcrosslinkers added to the reaction mix is such that the average numberof connections to the amine monomers (NC) is between about 2.05 andabout 6, or between about 2.2 and about 4.5. In some embodiments, thepolymer is further produced by a process where the target anion ispresent during the crosslinking reaction, for example by: a) adding theamine monomer as a free base and adding the target anion in its acidform; b) adding a crosslinker; c) carrying out the crosslinkingreaction; and d) washing out the target ion.

In some embodiments, the invention provides an anion-binding polymerwhere the polymer binds a target anion (e.g., phosphate or oxalate), andwhere the polymer is characterized by at least two of the followingfeatures: a) a swelling ratio of less than about 5; b) a gel pore volumedistribution measured in a physiological medium characterized by afraction of said pore volume accessible to non-interacting solutes, ofmolecular weight greater than about twice the MW of the target anion, ofless than about 20% of the weight of the gel; and c) an ion-bindinginterference for the target anion lower than about 60% when measured ina gastrointestinal simulant, relative to a non-interfering buffer, wherethe polymer contains one or more amine monomers and one or morecrosslinkers, and where the polymer is produced by a process including:a) forming soluble prepolymer by adding the entire amine monomercomponent and then continuously adding continuously a fraction of thecrosslinker to forming a syrup; b) emulsifying the syrup in oil; and c)adding the remaining fraction of crosslinker to form crosslinked beads.

In some embodiments, the invention provides an anion-binding polymerwhere the polymer binds a target anion (e.g., phosphate or oxalate), andwhere the polymer is characterized by at least two of the followingfeatures: a) a swelling ratio of less than about 5; b) a gel pore volumedistribution measured in a physiological medium characterized by afraction of said pore volume accessible to non-interacting solutes, ofmolecular weight greater than about twice the MW of the target anion, ofless than about 20% of the weight of the gel; and c) an ion-bindinginterference for the target anion lower than about 60% when measured ina gastrointestinal simulant, relative to a non-interfering buffer, wherethe polymer contains one or more amine monomers and one or morecrosslinkers, and where the polymer is produced by a process including:a) carrying out a first reaction between an amine monomer and acrosslinker to form a gel; then b) reacting the gel with anaminoalkylhalide, where the amine alkyl groups are chemically attachedto the gel through halide substitution by the amine functional gels.

In some embodiments, the invention provides a phosphate-binding polymercontaining one or more amine monomers and one or more crosslinkers wherethe polymer is produced by a process where the total content ofcrosslinkers added to the reaction mix is such that the average numberof connections to the amine monomers is between 2.2 and 4.5.

In some of these embodiments, the amine monomer is selected from thegroup consisting of 1,3 diamino propane, and N,N,N′,N′-tetrakis(3-aminopropyl) 1,4 diaminobutane, and wherein the crosslinker isselected from the group consisting of 1,3 dichloropropane andepichlorohydrin. In embodiments, the invention provides an ion-bindingpolymer comprising N,N,N′,N′-tetrakis (3-aminopropyl) 1,4 diaminobutanecrosslinked by epichlorohydrin wherein the polymer is produced by aprocess wherein the ratio of the initial concentration of N,N′ tetrakis(3-aminopropyl 1,4 diaminobutane to water is about 1:3 to about 4:1, orabout 1.5:1 to about 4:1.

In some embodiments, the invention provides a phosphate-binding polymercontaining N,N,N′,N′-tetrakis (3-aminopropyl) 1,4 diaminobutane monomersand epichiorohydrin crosslinker, wherein the polymer is produced by aprocess in which the total epichlorohydrin crosslinker added to thereaction mix is about 200% to about 300 mol %, or about 230 to about 270mol %, or about 250 mol % of the total N,N,N′,N′-tetrakis(3-aminopropyl) 1,4 diaminobutane content. In some of these embodiments,the polyniei is produced by a process in which the ratio of monomers towater in the initial reaction mix is about 3:1 to about 1:1, or about1.73 by weight. In some embodiments, the polymer is in the form ofspherical beads.

In some embodiments, the invention provides a phosphate-binding polymercomprising polyallylamine monomers and epichlorohydrin crosslinker,wherein the polymer is produced by initially dissolving thepolyallylamine monomers in water at a monomer:water ratio of about 3:1to about 1:3. In some of these embodiments, the total epichlorohydrincrosslinker added to the reaction mix is about 10 mol % of the totalpolyallylamine content.

In some embodiments, the invention provides a phosphate-binding polymercomprising a prepolymer comprising 1,3 diamino propane and 1,3dichloropropane crosslinker in a 1:1 molar ratio, wherein the prepolymeris further crosslinked by epichlorohydrin crosslinker, and wherein thetotal epichlorohydrin crosslinker added to the reaction mix is about 200mol % of the total prepolymer, and wherein the prepolymer:water ratio inthe reaction mix is about 1.1:1 to about 1.7:1.

The invention further provides compositions containing any of the abovepolymers where the polymer is in the form of particles, and where thepolymeric particles are encased in an outer shell.

In another aspect, the invention provides pharmaceutical compositions.In one embodiment, the pharmaceutical composition contains a polymer ofthe invention and a pharmaceutically acceptable excipient. In someembodiments, the composition is a liquid formulation in which thepolymer is dispersed in a liquid vehicle of water and suitableexcipients. In some embodiments, the invention provides a pharmaceuticalcomposition comprising an anion-binding polymer that binds a targetanion, and one or more suitable pharmaceutical excipients, where thecomposition is in the form of a chewable or mouth-disintegrating tablet,and where the polymer has a swelling ratio while transiting the oralcavity and in the esophagus of less than about 5, or less than about2.8, or less than about 2.7, or less than about 2.6, or preferably lessthan about 2.5. In some embodiments the chewable tablet contains polymerwhere the polymer has a transition temperature greater than about 50° C.

In some embodiments the chewable tablet contains a pharmaceuticalexcipient selected from the group consisting of sucrose, mannitol,xylitol, maltodextrin, fructose, sorbitol, and combinations thereof, andis produced by a process where the polymer is pre-formulated with theexcipient to form a solid solution. In some embodiments the target anionof the polymer is phosphate. In some embodiments the polymer binds atarget ion in vivo with a binding capacity of greater than 0.5 mmol/gr.In some embodiments the anion-binding polymer is more than about 50% ofthe weight of the tablet. In some embodiments, the tablet is ofcylindrical shape with a diameter of about 22 mm and a height of about 4mm and the anion binding polymer comprises more than about 1.6 gm of thetotal weight of the tablet. In some of the chewable tablets of theinvention, the excipients are chosen from the group consisting ofsweetening agents, binders, lubricants, and disintegrants. Optionally,the polymer is present as particles of less than about 40 um meandiameter. In some of these embodiments, the sweetening agent is selectedfrom the group consisting of sucrose, mannitol, xylitol, maltodextrin,fructose, and sorbitol, and combinations thereof.

In a further aspect, the invention provides a method of measuring targetion binding interference for an ion-binding polymer that binds a targetion by: a) adding the ion binding polymer to a non-interfering buffercontaining the target ion and measuring the target ion binding capacityof the polymer; b) artificially digesting a standardized meal withmammalian GI enzymes and/or aspirating chyme from the uppergastrointestinal tract of mammals having taken said standardized meal;wherein the standardized meal contains the target ion; c) adding the ionbinding polymer and measuring the target ion binding capacity from thetarget ion concentration decrease before and after the addition oftarget ion; and d) calculating the degree of interference in binding asthe fractional decrease in binding capacity for the target ion,expressed as a percent, observed between the binding measurement in anon interfering buffer, and in the digested meal or ex-vivo aspirates,at the same ion concentration in equilibrium.

In yet a further aspect, the invention provides a method of selecting anion-binding polymer, said polymer comprising monomer and crosslinker,wherein said polymer possesses at least one of the features: a) aswelling ratio of less than about 5; b) a gel pore volume distributionmeasured in a physiological medium characterized by a fraction of saidpore volume accessible to non-interacting solutes, of molecular weightgreater than about twice the MW of the target anion, of less than about20% of the weight of the gel; and c) an ion-binding interference for thetarget anion lower than about 60% when measured in a gastrointestinalsimulant, relative to a non-interfering buffer that includes the stepsof: i) varying the following composition and process variables: 1) theratio of crosslinker to monomer; 2) the ratio of (monomer+crosslinker)to solvent in the reaction medium; 3) the net charge of the polymer atphysiological pH and tonicity; and/or 4) the hydrophilic/hydrophobicbalance of the backbone polymer; ii) evaluating the swellability,porosity, and ion binding interference of the resulting polymer; andiii) selecting a polymer that possesses at least one of said features.In another aspect, the invention provides a method for improving thetherapeutic properties and/or suitability for administration and/orpharmaceutical properties of a polyamine polymer comprising at least oneof the following steps: a) crosslinking said polymer with a crosslinker,such that the average number of connection to the polyamine monomer isbetween about 2.05 and about 6; and/or b) producing said polymer by aprocess wherein the polyamine is initially present in water at a ratioof polyamine:water of from about 3:1 to about 1:3.

In another aspect, the invention provides a method of making ananion-binding polymer that binds a target anion, comprising combining anamine monomer with a crosslinker by a heterogeneous process, wherein thephosphate-binding polymer is characterized by at least two of thefollowing features: a) a swelling ratio of less than about 5; b) lessthan about 20% of the weight of the polymer accessible tonon-interacting solutes of molecular weight greater than about twice theMW of the target anion, wherein said percentage is measured in aphysiological medium, and c) an ion-binding interference for the targetanion lower than about 60% when measured in a gastrointestinal simulant,relative to a non-interfering buffer. In some embodiments the aminemonomer is a polyallylamine. In some embodiments the crosslinker isepichlorohydrin.

In another aspect the invention provides an anion-binding polymer thatbinds a target ion, wherein the polymer is produced by a processcomprising crosslinking a polyallylamine by a heterogeneous process, andwherein said polymer is characterized by at least two of the followingfeatures: a) a swelling ratio of less than about 5; b) less than about20% of the weight of the polymer accessible to non-interacting solutesof molecular weight greater than about twice the MW of the target anion,wherein said percentage is measured in a physiological medium, and c) anion-binding interference for the target anion lower than about 60% whenmeasured in a gastrointestinal simulant, relative to a non-interferingbuffer. In one embodiment, the polyallyamine is crosslinked byepichlorohydrin.

In another aspect, the invention provides a method of removing an anionfrom an animal by administering an effective amount of a polymer of theinvention to the animal. In some embodiments, the polymer is ananion-binding polymer where the polymer binds a target anion (e.g.,phosphate or oxalate), and where the polymer is characterized by atleast two of the following features: a) a swelling ratio of less thanabout 5; b) a gel pore volume distribution measured in a physiologicalmedium characterized by a fraction of said pore volume accessible tonon-interacting solutes, of molecular weight greater than about twicethe MW of the target anion, of less than about 20% of the weight of thegel; and c) an ion-binding interference for the target anion lower thanabout 60% when measured in a gastrointestinal simulant, relative to anon-interfering buffer. In some embodiments, the target anion of thepolymer is phosphate; in some embodiments the phosphate is removed fromthe gastrointestinal tract; in some embodiments the method ofadministration is oral. In some embodiments, the animal is afflictedwith at least one disease selected from the group consisting ofhyperphosphatemia, hypocalcemia, hyperthyroidism, depressed renalsynthesis of calcitriol, tetany due to hypocalcemia, renalinsufficiency, ectopic calcification in soft tissues, and ESRD. In someembodiments, the animal is a human.

In some embodiments, the polymer is co-administered with at least one ofproton pump inhibitor, calcimimetic, vitamin and analogs thereof, orphosphate binder, e.g., a phosphate binder that is aluminum carbonate,calcium carbonate, calcium acetate, lanthanum carbonate, or SEVELAMERhydrochloride.

INCORPORATION BY REFERENCE

All publications and patent applications mentioned in this specificationare herein incorporated by reference to the same extent as if eachindividual publication or patent application was specifically andindividually indicated to be incorporated by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph illustrating the determination of binding interferenceby comparing an isotherm of binding of target ion by a polymer in anon-interfering buffer to binding of target ion by the polymer in aninterfering medium (e.g., gastrointestinal simulant or ex vivoaspirate).

FIG. 2 is a graph illustrating non-accessible volume of gel versus probesolute radius.

FIG. 3 is a graph illustrating the determination of binding interferencefor a phosphate-binding polymer (EC172A).

FIG. 4 is a graph illustrating the determination of binding interferencefor a phosphate-binding polymer (RENAGEL).

FIG. 5 is a plot of nonaccessible volume versus probe molecular weight,for non-interacting probes, illustrating the difference between aphosphate-binding polymer of the invention (EC 172A) and acommercially-available phosphate binder (RENAGEL).

FIG. 6 is a plot of nonaccessible volume versus probe radius, fornon-interacting probes illustrating the difference between aphosphate-binding polymer of the invention (EC 172A) and acommercially-available phosphate binder (RENAGEL).

FIG. 7 is a graph illustrating the change in binding capacity withmodification of FR-005-144 with chloropropylamine, hydrochloride.

The novel features of the invention are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present invention will be obtained by reference to thefollowing detailed description that sets forth illustrative embodiments,in which the principles of the invention are utilized, and theaccompanying drawings of which:

DETAILED DESCRIPTION OF THE INVENTION

I. Introduction

One aspect of the invention provides anion binding polymeric materialsthat have one or more of the characteristics of low swelling, high ionbinding in vivo, low interference from interfering ions, and/or specificporosity. Another aspect of the invention provides pharmaceuticalcompositions of the anion-binding polymers, where the pharmaceuticalcomposition is a chewable tablet or a liquid formulation. A furtheraspect of the invention provides methods of making or improvinganion-binding polymers so that they have one or more of thecharacteristics of low swelling, high ion binding in vivo, lowinterference from interfering ions, and/or specific porosity. A yetfurther aspect of the invention is methods of using the anion-bindingpolymers of the invention to treat conditions in which an ion is inexcess. In a preferred embodiment, the anion-binding polymers are usedto remove target anions from the GI tract. Examples of target anionsthat can be removed from the GI tract include, but are not limited to,phosphate and oxalate. In another preferred embodiment, the compositionsdescribed herein are used in the treatment of hyperphosphatemia,hypocalcemia, hyperparathyroidism, depressed renal synthesis ofcalcitriol, tetany due to hypocalcemia, renal insufficiency, ecotopiccalcification in soft tissues, chronic renal insufficiency, and anabolicmetabolism.

II. Polymers

The polymers of the invention are characterized by their ability to bindions. Preferably the polymers of the invention bind anions, morepreferably they bind phosphate and/or oxalate, and most preferably theybind phosphate ions. For illustration, anion-binding polymers andespecially phosphate-binding polymers will be described; however, it isunderstood that this description applies equally, with appropriatemodifications that will be apparent to those of skill in the art, to allions and solutes. As used herein, a polymer “binds” an ion, e.g. ananion, or is an “ion-binding” polymer (e.g., a “phosphate-binding”polymer) when it associates with the ion, generally though notnecessarily in a noncovalent manner, with sufficient associationstrength that at least a portion of the ion remains bound under the invitro or in vivo conditions in which the polymer is used for sufficienttime to effect a removal of the ion from solution or from the body. A“target ion” is an ion to which the polymer binds, and usually refers tothe major ion bound by the polymer, or the ion whose binding to thepolymer is thought to produce the therapeutic effect of the polymer. Apolymer may have more than one target ion. “Binding” of an anion, ismore than minimal binding, i.e., at least about 0.01 mmole of anion/gmof polymer, more preferably at least about 0.05 mmole of anion/gm ofpolymer, even more preferably at least about 0.1 mmole of anion/gm ofpolymer, and most preferably at least about 0.5 mmole of anion/gm ofpolymer. The invention provides polymers that are characterized by theirselective binding of anions; for example, in some embodiments, polymersof the invention bind bile acids with a binding capacity of less thanabout 2 mmol/gm, preferably less than about 1 mmol/gm, more preferablyless than about 0.5 mmol/gm, even more preferably less than about 0.3mmol/gm, and most preferably less than about 0.1 mmol/gm. In someembodiments, polymers of the invention bind citrate with a bindingcapacity of less than about 2 mmol/gm, preferably less than about 1mmol/gm, more preferably less than about 0.5 mmol/gm, even morepreferably less than about 0.3 mmol/gm, and most preferably less thanabout 0.1 mmol/gm.

A. Characteristics

The polymers of the invention are characterized by one or more of thefollowing features: 1) low swelling ratio; 2) low binding interferenceunder physiological conditions; 3) a porosity appropriate to bind thetarget anion, and to exclude interfering solutes; 4) an in vivo bindingcapacity for the target anion, sufficient to be effective in therapeuticuses. In some embodiments, the polymer is an anion-binding polymer(e.g., a polymer that binds phosphate and/or oxalate), where the polymeris characterized by at least two of the following features: 1) aswelling ratio of less than about 5; 2) a gel pore volume distributionmeasured in a physiological medium characterized by less than about 20%of said pore volume accessible to non-interacting water soluble solutesof molecular weight greater than about twice the MW of the target anion;and 3) an ion-binding interference for the target anion of said polymerlower than about 60% when measured in a gastrointestinal simulant,relative to a non-interfering buffer. In some embodiments, the polymeris a phosphate-binding polymer characterized by at least one of thefollowing features: 1) a swelling ratio of less than about 5, preferablyless than about 2.5; 2) a gel pore volume distribution measured in aphysiological medium characterized by less than about 20% of said porevolume accessible to non-interacting solutes of molecular weight greaterthan about 200; and 3) an ion-binding interference for phosphate lowerthan about 60% when measured in a gastrointestinal simulant, relative toa non-interfering buffer. In some embodiments, the phosphate-bindingpolymer has a swelling ratio of less than about 2.8, or less than about2.7, or less than about 2.6. A “physiological medium” is a medium thatis isotonic and at neutral pH. In some embodiments the inventionprovides a phosphate-binding polymer characterized by a swelling ratio,measured in isotonic medium at neutral pH, of less than about 5,preferably less than about 2.5, optionally with a mean in vivo bindingcapacity for phosphate of greater than about 0.5 mole/gm. In someembodiments, the phosphate-binding polymer has a swelling ratio of lessthan about 2.8, or less than about 2.7, or less than about 2.6. In someembodiments, polymers of the invention bind bile acids with a bindingcapacity of less than about 2 mmol/gm, preferably less than about 1mmol/gm, more preferably less than about 0.5 mmol/gm, even morepreferably less than about 0.3 mmol/gm, and most preferably less thanabout 0.1 mmol/gm. In some embodiments, polymers of the invention bindcitrate with a binding capacity of less than about 2 mmol/gm, preferablyless than about 1 mmol/gm, more preferably less than about 0.5 mmol/gm,even more preferably less than about 0.3 mmol/gm, and most preferablyless than about 0.1 mmol/gm. Preferably, the polymers are composed ofamine monomers.

Generally, these features are achieved by manipulating one or morefactors in the production of the polymer.

1) Swelling ratio. Polymers of the invention are crosslinked materials,meaning that they do not dissolve in solvents, and, at most, swell insolvents.

The ratio of swelling in physiological isotonic buffer, representativeof the milieu of use, i.e. the gastrointestinal tract, is typically inthe range of about 1.2 to about 100, preferably about 2 to 20. In someembodiments, polymers of the invention have a swelling ratio of lessthan 5, or less than about 4, or less than about 3, or less than about2.8, or less than about 2.7, or less than about 2.6, or less than about2.5. As used herein, “swelling ratio” refers to the number of grams ofsolvent taken up by one gram of dried crosslinked polymer, whenequilibrated in an aqueous environment. When more than one measurementof swelling is taken for a given polymer, the mean of the measurementsis taken to be the swelling ratio.

Swelling ratios are measurable using a variety of methods: the mostpreferred is the gravimetric method, in which the dried polymer isweighed and added to an excess of liquid. In some cases the liquid maybe distilled water; preferably the liquid is an aqueous solution that isisotonic to plasma; most preferably the liquid is an aqueous solutionthat is isotonic to plasma and that is buffered to a neutral pH. Forexample, 0.9% NaCl solution may be used. Phosphate buffered saline (PBS)may also be used. The most preferred physiological medium for swellingmeasurements is 0.9% NaCl buffered with 30 mM MES to a pH of betweenabout 6.5 and 7.5. The dry polymer (e.g., a phosphate binding plymer)generally is used in fully protonated form with counterion, e.g.,chloride. The polymer is soaked in the liquid until equilibration. Thesoaked gel is then centrifuged, the supernatant decanted, and the wetgel weighed. Care should be taken not to centrifuge to too high g numberto avoid gel collapse. The swelling ratio (SR) is calculated as theweight of wet gel minus the weight of dry polymer, divided by the weightof dry polymer.

Another method is the dye method, in which a dye of very high molecularweight and known not to interact with the gel is prepared as an aqueoussolution and an aliquot of dry polymer is added to the solution. Theweight to weight ratio of solution to polymer is adjusted to be closeand slightly higher than the expected swelling ratio. As the dye is ofvery high molecular weight (e.g. greater than 200,000 g/mol), it doesnot permeate the gel while the water does, leading to an increase in theresulting dye concentration, from which the swelling ratio isdetermined. An example of useful dye is dextran modified withfluoresceine isothiocyanate (FITC).

The swelling ratio of a polymer depends on a number of variables such astemperature, ionic strength, polymer charge density, polymer-solventFlory-Huggins coefficient and crosslinking density. Because theion-binding polymers of the invention are mostly charged polymers (e.g.phosphate ion binding polyamine are protonated at intestinal pH), theirswelling behavior is typical of polyelectrolyte gels. Although swellingratio and pore size are somewhat related, i.e. large swelling ratio isusually accompanied by large pores, there is no theoretical basis toaccurately predict the exclusion limit of polyelectrolyte gels.

2) Binding interference. In some embodiments, polymers of the inventionhave a binding interference of less than about 70%, more preferably lessthan about 60%, even more preferably less than about 50%, morepreferably less than about 40%, even more preferably less than about30%, and most preferably less than about 20%, when measured in agastrointestinal (GI) simulant. Phosphate-binding polymers of theinvention exhibit a binding interference of less than about 70%, morepreferably less than about 60%, even more preferably less than about50%, more preferably less than about 40%, even more preferably less thanabout 30%, and most preferably less than about 20%, when measured in aGI simulant.

The “degree of interference in binding” or “binding interference,” asused herein, refers to the fractional decrease in binding capacity forthe target ion, expressed as a percent, observed between a bindingexperiment in a non interfering buffer, and in a gastrointestinal (GI)simulant, at the same concentration of target anion in equilibrium. A“non-interfering buffer,” as used herein, refers to a buffer that doesnot contain one or more solutes that interfere with the binding of thetarget ion, and that is buffered to the same pH as the GI simulant. Anon-interfering buffer is not necessarily free of all interferingsolutes, for example a non-interfering buffer may contain one or both ofthe ubiquitous gastrointestinal ions chloride and bicarbonate; ifpresent, these may be at their physiological concentration. An exampleof a non-interfering buffer is given in Example 1. A “GI simulant”refers herein to a preparation that is designed to mimic the milieu of aportion of the GI tract after ingestion of a meal, preferably theportion of the GI tract in which the polymer will be binding themajority of target ion. The GI simulant typically is prepared by themethod illustrated in Example 1. Target ion should be present in the GIsimulant at the same concentration(s) as used in non-interfering bufferstudies. The degree of interference is easily illustrated by plottingthe two corresponding binding isotherms, i.e. GI simulant and in anon-interfering buffer, as shown in FIG. 1. An example of determinationof binding interference using a GI simulant is given in Example 1.

It is also possible to measure binding interference by comparing bindingof target ion in digestive aspirate from subjects, preferably humansubjects, to binding of target ion in non-interfering buffer. If thismeasurement is done, aspirates should be obtained from a number ofsubjects and the mean interference taken as the binding interference.

It has been found that by carefully selecting the swelling ratio and/oradjusting the molecular weight exclusion limit of the gel, the bindingcapacity measured in a competitive mode (i.e. in vivo or in a GIstimulant) can be substantially increased compared with other gels withthe same polymer composition but otherwise non-optimized in gelporosity.

Strikingly, polymers in which crosslinking and/or entanglement wereincreased were found to have lower swelling than those with lowercrosslinking and/or entanglement, yet also had a binding capacity fortarget ion (e.g., phosphate) that was as great as or greater than thelower crosslinking and/or entanglement polymers. Not wishing to be boundby theory, it is hypothesized that polymers of the invention exert asieving effect and bind only solutes of a specific size in solution andexclude other larger species that would otherwise compete with bindingsites within the polymer. Larger molecular weight species include butare not limited to, inorganic and organic anions, oligopeptides,carbohydrates, bilirubins, lipid micelles and lipid vesicles.

3) Porosity. It has been found that it is possible to manipulate theprocess for producing a polymer so that the polymer more optimallyexhibits a porosity appropriate to bind the target ion (e.g., anions)for which the polymer is intended and to exclude interfering substances.

Pore size distribution of polymeric gels is obtained by various methodssuch as mercury porosimetry, nitrogen adsorption, differential scanningcalorimetry, or solute permeation partitioning techniques. The lattertechnique, solute permeation partitioning technique, is most preferredas it probes the gel in a fully hydrated state identical to oneprevailing in the milieu of use. The solute permeation technique is anindirect method introduced by Kuga (Kuga S. J, J. of Chromatography,1986, 206: 449-461) and consists of measuring the gel partitioning ofsolutes of known molecular weights. This method consists of three majorsteps (Kremer et al., Macromolecules, 1994, 27, 2965-73):

-   -   1. Solutions with dissolved solutes of known concentrations and        molecular sizes are brought into contact with the swollen gel.        The molecular sizes of the solutes must cover a substantial        range.    -   2. Diffusion of solutes into the gel is attained. Partitioning        of a particular solute depends on both the size of the solute        and the size distribution of the gel pores.    -   3. The gel is separated from its surrounding solution, and        subsequent concentration measurement of solutes in the        surrounding solution are made. The decrease of each solute        concentration relative to its initial stock solute concentration        is used for calculating the gel pore size distribution.

To isolate the size exclusion effects from molecularattraction/repulsion effects, the solutes are selected from polymers oroligomers that have little or no interactions with the gel polymer;neutral hydrophilic polymers with narrow molecular weight distributionsuch as polyethyleneglycol, polyethylene oxide or dextran, are mostsuitable. Thus, unless otherwise indicated herein, volumes for exclusionof particular sizes of solutes (also referred to herein as “criticalpermeation volume”) refer to volumes measured using solutes withsubstantially no interaction with the polymer for which measurements aretaken.

Following the experimental protocol and data treatment given in Kremeret al., Macromolecules, 1994, 27, 2965-73, the pore size distributionmay be represented as shown in FIG. 2. In FIG. 2, the Y axis representsthe volume of the swollen gel not accessible to a solute of a givenmolecular size. In the example shown in the Figure, small molecules witha size smaller than 5 angstroms can permeate throughout the entire gel.At the other extreme, polymers with a hydrodynamic radius greater than1000 angstroms are totally excluded from the gel. In that case the nonaccessible volume and the volume of gel at equilibrium are the same.

Size and molecular weight of polymers are related through Mark-Houvinkequations which are tabulated for the polymer solutes used as molecularprobes. For example:

-   -   Radius (angstroms)=0.217M^(0.498) Dextran    -   Radius (angstroms)=0.271 M^(0.517) Polyethyleneglycol    -   Radius (angstroms)=0.166M^(0.573) Polyethyleneoxide

Small molecular weight probes can also be used:

-   -   Urea: Molecular radius 2.5 angstroms    -   Ethylene glycol: Molecular radius 2.8 angstroms    -   Glycerol: Molecular radius 3.1 angstroms    -   Glucose: Molecular radius 4.4 angstroms    -   Saccharose: Molecular radius 5.3 angstroms

Thus, the size for the solute may be converted to molecular weight andvice-versa.

The size of the solute does not equate the size of the pores; otherwisethis would mean that all the liquid existing in pores greater than themolecular size of the solute is available as accessible volume: this isincorrect because of the excluded volume effect, also known as the walleffect.

A straightforward manner to characterize the molecular exclusion limitis: (i) quantify the partitioning of molecular probes, (ii) calculatethe accessible volume (or weight) as described above and (iii) normalizeit to the total gel volume (or weight).

The desired molecular exclusion limit is achieved by manipulation ofproduction variables such as entanglement of polymer strands andconcentration of crosslinker (see below). In general, polymers areproduced to have a molecular exclusion limit that is based on the ion(e.g., anion) to be bound and the probable identity of the interferingsubstances that are wished to be excluded, as well as the tolerableamount of swelling for the intended use of the polymer. In someembodiments of the invention, the ion binding polymer displays a gelpore volume distribution (critical permeation volume) defined accordingto the protocol described above and measured in a physiological mediumof less than about 60%, less than about 40%, or less than about 20% ofthe polymer pore volume accessible to non-interacting solutes ofmolecular weight greater than about twice the MW of the target anion. Insome embodiments of the invention, the ion binding polymer displays agel pore volume distribution (critical permeation volume) of less thanabout 60%, less than about 40%, or less than about 20% of the polymerpore volume accessible to non-interacting solutes of molecular weightgreater than about 1.8-fold the MW of the target ion. In someembodiments of the invention, the ion binding polymer displays a gelpore volume distribution (critical permeation volume) of less than about60%, less than about 40%, or less than about 20% of the polymer porevolume accessible to non-interacting solutes of molecular weight greaterthan about 1.6-fold the MW of the target ion. In some embodiments of theinvention, the ion binding polymer displays a gel pore volumedistribution (critical permeation volume) of less than about 60%, lessthan about 40%, or less than about 20% of the polymer pore volumeaccessible to non-interacting solutes of molecular weight greater thanabout 1.4-fold the MW of the target ion. In some embodiments of theinvention, the ion binding polymer displays a gel pore volumedistribution (critical permeation volume) of less than about 60%, lessthan about 40%, or less than about 20% of the polymer pore volumeaccessible to non-interacting solutes of molecular weight greater thanabout 1.2-fold the MW of the target ion. In embodiments the inventionprovides a phosphate-binding polymer displaying a gel pore volumedistribution (critical permeation volume) defined according to theprotocol described above and measured in a physiological medium of lessthan about 60%, less than about 40%, or less than about 20% of thepolymer pore volume accessible to non-interacting solutes of molecularweight greater than about 200, more preferably greater than about 180,more preferably greater than about 160, more preferably greater thanabout 140, and most preferably greater than about 120.

4. Binding capacity. The polymers described herein exhibit ion bindingproperties, generally anion-binding properties. In preferredembodiments, the polymers exhibit phosphate binding properties. Ion(e.g., phosphate) binding capacity is a measure of the amount of aparticular ion an ion binder can bind in a given solution. For example,binding capacities of ion-binding polymers can be measured in vitro,e.g., in water or in saline solution, or in vivo, e.g., from ion (e.g.,phosphate) urinary excretion, or ex vivo, for example using aspirateliquids, e.g., chyme obtained from lab animals, patients or volunteers.Measurements can be made in a solution containing only the target ion,or at least no other competing solutes that compete with target ions forbinding to the polymer. In these cases, a non interfering buffer wouldbe used. Alternatively, measurements can be made in the presence ofother competing solutes, e.g., other ions or metabolites, that competewith target ions for binding to the resin.

Ion binding capacity for a polymer can be calculated asV*(C_(start)-C_(cq))/P, expressed in mmol/gr, where V is the fixedvolume of the solution used, in L; C_(start) is the initial target ionconcentration of the solution in mM; C_(eq) is the equilibrium targetion concentration in the solution in mM, after a weight P, in grams, ofpolymer is added and equilibration allowed.

In some embodiments the polymer binds phosphate. For in vivo use, e.g.,in treating hyperphosphatemia, it is desirable that the polymer have ahigh phosphate binding capacity. In vitro measurements of bindingcapacity do not necessarily translate into in vivo binding capacities.Hence, it is useful to define binding capacity in terms of both in vitroand in vivo capacity.

The in vitro phosphate binding capacity of the polymers of the inventionin a non-interfering buffer can be greater than about 0.5, 1.0, 1.5,2.0, 2.5, 3.0, 3.5, 4.0, 5.0, 6.0, 8.0, or 10.0 mmol/gr. In someembodiments, the in vitro phosphate binding capacity of the polymers ofthe invention for target ion is greater than about 0.5 mmol/gr,preferably greater than about 2.5 mmol/gr, even more preferably greaterthan about 3 mmol/gr, even more preferably greater than about 4 mmol/gr,and yet even more preferably greater than about 6 mmol/gr. In someembodiments, the phosphate binding capacity can range from about 0.5mmol/gr to about 10 mmol/gr, preferably from about 2.5 mmol/gr to about8 mmol/gr, and even more preferably from about 3 mmol/gr to about 6mmol/gr. Several techniques are known in the art to determine thephosphate binding capacity. The in vitro phosphate binding capacity ofthe polymers of the invention is measured as described in Example 1 formeasurement of binding capacity in a non-interfering buffer.

In some embodiments, the mean ex vivo phosphate binding capacity ofphosphate-binding polymers of the invention, measured in digestiveaspirates from human subjects, is greater than about 0.2, 0.3, 0.4, 0.5,0.6, 0.7, 0.8, 0.9, 1.0, 1.2, 1.4, 1.6, 1.8, 2.0, 2.5, 3.0, 4.0, 5.0, or6.0 mmol/gr. Ex vivo aspirates are obtained as described in Example 1,from normal subjects, and binding is measured as for a non-interferingbuffer. Mean values are taken from about 5-15, or about 15-30, or about30-60 subjects. In some embodiments, measurements are taken from 6-12subjects.

As used herein, “mean in vivo phosphate binding capacity” refers to thebinding capacity of a polymer as measured in normal human subjectsunless otherwise specified, and where the phosphate binding of thepolymer is measured by the decrease in the phosphate urinary excretioncombined with measurement of the phosphate excreted in the feces as freeand polymer-bound phosphate (see below). Mean values are taken fromabout 5-15, or about 15-30, or about 30-60 subjects. In someembodiments, measurements are taken from 6-12 subjects. In someembodiments, the mean in vivo phosphate binding capacity of the polymersof the invention, preferably measured in human subjects, is at leastabout 0.3 mmol/gr, at least about 0.5 mmol/gr, at least about 0.8mmol/gr, at least about 1.0 mmol/gr, at least about 1.5 mmol/gr, atleast about 2.0 mmol/gr, at least about 3.0 mmol/gr, at least about 4.0mmol/gr, at least about 5.0 mmol/gr, or at least about 6.0 mmol/gr.

The in vivo binding capacity of the polymer can preferably be determinedby measuring the balance of the target ion (e.g. phosphate ion) inmammals, preferably in humans: subjects are given a meal with acontrolled content of phosphate and binding polymer, and are monitoredfor phosphate intake and phosphate excreted in the feces and in urine.The study comprises a washout period followed by a period where subjectstake a daily dose, preferably t.i.d., of phosphate binder, followed byseveral days without treatment to observe the return to baseline. Thedepletion of phosphate in urine usually matches the increase ofphosphate in the feces. The moles of phosphate excreted in the fecesminus the baseline, divided by the weight of binder administeredprovides a measure of the in vivo binding capacity. Unless otherwiseindicated, “in vivo” measurements referred to herein utilize the aboveprotocol. Another method consists of measuring the phosphate binding invivo and in situ following the protocol indicated in Example 1, whereinmammals are intubated by a double lumen tube to retrieve the chyme at acertain location of the small intestine. A meal with a given phosphatecontent is given together with a known content of phosphate binder and amarker. The marker can be a dye or a non absorbable polymer (e.g.polyethylene glycol), which is then titrated in the chyme to determinethe dilution occurring during the digestion process. The actualconcentration of binder is then calculated from the initialconcentration in the meal and the dilution ratio measured from themarker experiment. The total phosphate is analyzed on the chyme sample.The “soluble” phosphate is measured by spinning down the sample anddecanting the supernatant and assaying for phosphate. The “bound”phosphate is obtained by difference between the total and the solublephosphate. Two series of experiments are made on a group of subjects(6-12) which alternatively take a placebo (microcrystalline cellulose)or the drug: The binding capacity is obtained by measuring the increasein “bound” phosphate between the two sets of experiments, i.e. with andwithout drug administration, and dividing by the concentration ofbinder. Calculation can be made either on a subject basis or on pergroup basis.

B. Preparation of Polymers

The polymers of the invention are prepared by methods known to thoseskilled in the art; for example: ion binding monomers or theirprecursors can be copolymerized in the presence of a crosslinker; apreformed ion binding polymer is subsequently crosslinked through achemical reaction or irradiation; or a polymer precursor is firstcrosslinked and further reacted to generate ion binding functionalgroups on the polymer.

The polymers are obtained by direct or inverse suspension, emulsion,precipitation techniques, polymerization in aerosol or using bulkpolymerization/crosslinking methods and size reduction processes such asextrusion and grinding. Processes can be carried out as batch,semi-continuous and continuous processes.

The swelling ratio, binding interference, binding capacity, and MWexclusion limit are affected by at least the following composition andprocess variables:

-   -   1—Concentration of the chemical crosslinks of the polymer        chains.    -   2—The (monomer+crosslinker) to solvent ratio in the crosslinking        reaction.    -   3—The net charge of the polymer (at the physiological pH and        tonicity of the milieu in which it will be used).    -   4—The hydrophilic/hydrophobic balance of the backbone polymer.    -   5—The presence or absence of a core-shell structure, where the        shell component restricts the extent of swelling of the core        material.

In the following, the preferred operating ranges for composition andprocess variables are exemplified with crosslinked polyamine materialswith phosphate binding properties. It will be understood that these areexemplary conditions only, and that the methods described herein may beused in the selection and production of polymers that bind a wide rangeof solutes, as will be apparent to one of skill in the art.

1) Concentration of the chemical crosslinks of the polymer chains. Theconcentration of chemical crosslinks is one important feature thatcontrols the swelling properties and pore distribution of the polymer.One convenient way to describe the polymers of the invention is todefine a amine repeat unit and its average number of connections to therest of the polymer. “A” is defined as the amine repeat unit and “NC” isthe average number of connections from A; NC can be 2, 3, 4 and higher.In order to form an insoluble gel, NC generally should be greater than2.

NC values can be then translated into amine to crosslinkerstoichiometric ratios by the following equations:

For low molecular weight monomers, e.g., N,N,N′,N′-tetrakis(3-aminopropyl) 1,4 diaminobutane or 1,3 diamino propane, NC=B*Fb/A,where B is the number of moles of crosslinker, Fb is the number ofgroups in B reacting with A to establish a covalent bond, and A is thenumber of moles of amine.

When the amine material is of high molecular weight and derives from thepolymerization of an amine monomer, such as vinylamine,polyethyleneimine, polyvinylamine, or polyallylamine, the expression ischanged to account for the 2 connections linking the monomer repeatswithin the polymer backbone. NC then becomes: NC=(2*A+B*Fb)/A.

Conversely, the mole ratio of crosslinker to amine can be computed fromthe desired NC value by manipulating the equations above:

Low molecular weight amine:B/A=NC*Fb

High molecular weight amine:B/A=(NC−2)*Fb

The table below shows some conversion examples between NC and the actualcrosslinker to amine ratio, by mole, wherein the amine is either a highmolecular weight material or a small molecule, and where the crosslinkermaterial is either di or tri functional.

Desired Mole ratio Equation Amine material Type of amine Crosslinkers FbNC B/A used Polyallylamine High Mw epichlorhydrine 2 2.2 0.10 bPolyvinylamine High Mw 1,3 dichloropropane 2 2.5 0.25 bpolyethyleneimine High Mw N-tris(2chloro ethyl)amine 3 2.2 0.07 b 1,3diaminopropane Low Mw amine 1,3 dichloropropane 2 2 1.00 aN,N,N′,N′-tetrakis (3-aminopropyl) 1,4 diaminobutane Low Mw amineepichlorhydrine 2 4 2.00 a N,N,N′,N′-tetrakis (3-aminopropyl) 1,4diaminobutane Low Mw amine N-tris(2chloro ethyl)amine 3 4 1.33 a (a):B/A = NC/Fb (b): B/A = (NC − 2)/Fb

Surprisingly it was found that the binding selectivity, which reflectsthe in vivo efficacy, went through an optimum with respect to NC: in thelow range of NC values, the material tended to swell considerably andcorollary showed a lot of binding interference in a GI stimulant. In thehigh range however, the material had substantially low intrinsic bindingcapacity which obviously lowered the overall performance in vivo. Theoptimal NC values was found to lie between 2.05 and 5 depending upon theamine/crosslinker systems.

However the optimal range for giving the desired combination ofcharacteristics in the final polymer depends on the specific monomer andcrosslinker used, as well as other conditions used in the productionprocess, such as initial concentration of the monomer in the reactionmedium, and is a matter of routine experimentation.

In some embodiments, the ratio of crosslinker to total amine groups ofthe monomers in the polymer is greater than 50 mol %, 60 mol %, 70 mol%, 80 mol %, or 90 mol %.

In some embodiments of the invention providing a phosphate-bindingpolymer containing one or more low molecular weight amine monomers andone or more crosslinkers, NC is more than about 2, or more than about 3,or more than about 4. In some embodiments polymers are constructed fromN,N,N′,N′-tetrakis (3-aminopropyl) 1,4 diaminobutane monomers (low MWmonomers) crosslinked by epichlorohydrin (Fb=2), where B/A is from about2.0 (mol/mol) to about 3.0 (mol/mol) (i.e, NC is from about 4 to about6), or from about 2.3 (mol/mol) to about 2.7 (mol/mol) (i.e, NC is fromabout 4.6 to about 5.4), or about 2.5 (mol/mol) (i.e, NC is about 5.0).In some embodiments polymers are constructed from N,N,N′,N′-tetrakis(3-aminopropyl) 1,4 diaminobutane monomers crosslinked byepichlorohydrin, where the initial ratio of monomer to water is fromabout 3:1 w/w to 1:3 w/w, or from about 1.5:1 to about 2:1 w/w, or about1:1, or about 3:1, where B/A is from about 2.0 (mol/mol) to about 3.0(mol/mol) (i.e, NC is from about 4 to about 6), or from about 2.3(mol/mol) to about 2.7 (mol/mol) (i.e, NC is from about 4.6 to about5.4), or about 2.5 (mol/mol) (i.e, NC is about 5.0).

2) The (monomer+crosslinker) to solvent ratio in the crosslinkingreaction. High ratios of (monomer+crosslinker) to solvent favor denselycrosslinked materials when all other conditions are kept constant. Forinstance, when a high molecular weight amine is used and when the chainlength and the polymer concentration are large enough, chainentanglements are produced that generate many crosslinking nodes oncethe structure is chemically crosslinked. More generally, for both highand low molecular weight amines, high (monomer+crosslinker) to solventratio tends to minimize the extent of side reactions leading to geldefects (e.g. intrachain crosslinking leading to cyclic structures,incomplete crosslinking reaction leading to dangling ends).

This condition is determined primarily by the concentrations in thereaction medium of both the monomer (e.g., amine) and the crosslinker.In some embodiments of the invention, the concentration of monomer andcrosslinker in the reaction medium is greater than about 20 wt %,preferably greater than 40 wt %, more preferably greater than 60% wt %.In some embodiments, a (monomer+crosslinker):solvent (e.g., water) ratioof between about 3:1 to about 1:3 (w/w) is used. In some embodiments, a(monomer+crosslinker):solvent (e.g., water) ratio of between about 3:1to about 1:1 (w/w) is used In some embodiments, a(monomer+crosslinker):solvent (e.g, water) ratio of about 3:1, or about2.5:1, 01 about 2.0:1, or about 1.5:1, or about 1:1 (w/w) is used. Thecrosslinker may be added at various times, depending on thepolymerization procedure in some embodiments, the initialmonomer:solvent ratio (before addition of era sslinker) is between about4:1 to about 1:1, or between about 3:1 to about 1:1; crosslinker is thenadded to comprise between about 100 mol % to about 400 mol % of theinitial monomer content, or between about 200 mol % to about 300 mol %of the initial monomer content In some embodiments, the monomer isN,N,N′,N′-tetrakis (3-aminopropyl) 1,4 diaminobutane monomers and thecrosslinker is epichiorohydrin, and the initial monomer:water ratio isbetween about 4:1 to 1:1, or between about 3:1 to about 1:1, or about1.7, or about 1.73 by weight; and the crosslinker is added to betweenabout 200 mol % to about 300 mol % of the monomer content, or about 230mol % to about 270 mol %, or about 250 mol %.

In some embodiments, e.g. embodiments in which the monomer is apolyallyamine, the amount of monomer is much greater than the amount ofcrosslinker (e.g., ten-fold crosslinker on a molar basis and evengreater on a weight basis), and the above ratios may be expressed asmonomer:solvent ratios, ignoring crosslinker. In some embodiments, themonomer (e.g., polyallylamine) is present at a monomer:solvent ratio ofabout 3:1 to about 1:3. In some embodiments, the monomer ispolyallylamine and the crosslinker is epichlorohydrin, where thepolyallylamine is present at monomer:water ratio of about 3:1 to about1:3, and epichlorhydrin is added to the reaction mix to about 10 mol %of the total polyallylamine content.

When possible solvent free process are even more preferred: in oneembodiment the amine and the crosslinker are quickly mixed andsubsequently dispersed neat in a continuous phase, e.g. water. Thecrosslinking reaction is taking place within the dispersed droplets andrecovered as beads.

3) The net charge of the polymer (at the physiological pH and tonicity).The net charge of the polymer is given by the mole content of theion-binding, its intrinsic charge and degree of ionization atphysiological pH. The charge density is preferably in the range of 3 to20 mmol/gr, preferably 6 to 15 mmol/gr.

4) The hydrophilic/hydrophobic balance of the backbone polymer. Thehydrophilic/hydrophobic balance of the polymer allows one to controlsomewhat independently the chemical crosslinking density and theswelling ratio. The swelling ratio is very sensitive to the polymersolvent interaction parameter χij as described in the Flory-Hugginsformalism (Flory P. J. “Principles of Polymer Chemistry, Cornell IthacaPub. 1953)). Increasing χij values up to 0.4 and above create a poorsolvent conditions for the polymer, which then tries to minimize monomerto solvent (water) interaction and consequently swells much less. Thiscan be achieved by incorporating hydrophobic moieties in the gel, suchas long chain hydrophob, (poly)aromatic substituents, or fluorinatedgroups. When this strategy is chosen to control the extent of swellingand consequently the exclusion limit of the gels, the level ofhydrophobic monomers and crosslinkers is between about 0.5 mol % toabout 50 mol %, preferably between about 20% and 50%.

In preferred methods, the absolute hydrophobicity is quantified by theabsolute difference in the log P of the monomers. Quantitatively, thehydrophobic/hydrophilic nature of the monomers may be determinedaccording to the log P of the particular monomers, which is sometimesreferred to as the octanol-water partition coefficient. Log P values arewell known and are determined according to a standard test thatdetermines the concentration of monomer in a water/1-octanol separatedmixture. In particular, computer programs are commercially available aswell as on the internet that will estimate the log P values forparticular monomers. Some of the log P values in this application wereestimated from the web sitehttp://esc.syrres.com/interkow/kowdemo.htm,which provides an estimated log P value for molecules by simplyinserting the CAS registry number or a chemical notation. Hydrophobicmonomers typically will have a log P value above zero and hydrophilicmonomers typically will have a log P value close to or below zero.Generally, the log P of the hydrophobic monomers for the purposes ofthis invention should be at least about 0.5, more preferably at leastabout 0.75, even more preferably at least about 1.0, still morepreferable at least about 1.5 and most preferably at least about 2.

5) The presence of a core-shell structure where the shell componentrestricts the extent of swelling of the core material. Gel particleswith a core-shell morphologies are useful in the context of theinvention: the shell material can limit the swelling, hence limit theexclusion limit, by imposing a mechanical resistance on the swellingpressure stemming from the core material, which would otherwise swell toa much higher extent. The shell material can be of the same compositionthan the core, but with a higher crosslink density. The design of suchcore-shell materials and method of making thereof can be found in U.S.patent application Ser. No. 10/814,789.

The shell material can be chemically anchored to the core material orphysically coated. In the former case, the shell can be grown on thecore component through chemical means, for example by: chemical graftingof shell polymer to the core using living polymerization from activesites anchored onto the core polymer; interfacial reaction, i.e., achemical reaction located at the core particle surface, such asinterfacial polycondensation; and using block copolymers as suspendingagents during the core particle synthesis.

The interfacial reaction and use of block polymers are preferredtechniques when chemical methods are used. In the interfacial reactionpathway, typically, the periphery of the core particle is chemicallymodified by reacting small molecules or macromolecules on the coreinterface. For example, an amine containing ion-binding core particle isreacted with a polymer containing amine reactive groups such as epoxy,isocyanate, activated esters, halide groups to form a crosslinked shellaround the core.

In another embodiment, the shell is first prepared using interfacialpolycondensation or solvent coacervation to produce capsules. Theinterior of the capsule is then filled up with core-forming precursorsto build the core within the shell capsule.

In some embodiments, using the block copolymer approach, an amphiphilicblock copolymer can be used as a suspending agent to form the coreparticle in an inverse or direct suspension particle forming process.When an inverse water-in-oil suspension process is used, then the blockcopolymer comprises a first block soluble in the continuous oil phaseand another hydrophilic block contains functional groups that can reactwith the core polymer. When added to the aqueous phase, along withcore-forming precursor, and the oil phase, the block copolymer locatesto the water-in-oil interface and acts as a suspending agent. Thehydrophilic block reacts with the core material, or co-reacts with thecore-forming precursors. After the particles are isolated from the oilphase, the block copolymers form a thin shell covalently attached to thecore surface. The chemical nature and length of the blocks can be variedto vary the permeation characteristics of the shell towards solutes ofinterest.

When the shell material is physically adsorbed on the core material,well known techniques of microencapsulation such as solventcoacervation, fluidized bed spray coater, or multiemulsion processes canbe used. A preferred method of microencapsulation is the fluidized bedspray coater in the Wurster configuration. In yet another embodiment,the shell material is only acting temporarily by delaying the swellingof the core particle while in the mouth and esophagus, and optionallydisintegrate in the stomach or duodenum. The shell is then selected inorder to hinder the transport of water into the core particle, bycreating a layer of high hydrophobicity and very low liquid waterpermeability.

Thus, in one aspect the invention provides a method of selecting anion-binding polymer, where the polymer contains a monomer and acrosslinker, and where the polymer possesses at least one of thefeatures of a) a swelling ratio of less than about 5; b) a gel porevolume distribution measured in a physiological medium characterized bya fraction of said pore volume accessible to non-interacting solutes, ofmolecular weight greater than about twice the MW of the target anion, ofless than about 20% of the weight of the gel; and c) an ion-bindinginterference for the target anion lower than about 60% when measured ina gastrointestinal simulant, relative to a non-interfering buffer, by:

-   -   i) varying the following composition and process variables        -   1) the ratio of crosslinker to monomer;        -   2) the ratio of (monomer+crosslinker) to solvent in the            reaction medium;        -   3) the net charge of the polymer at physiological pH and            tonicity; and/or        -   4) the hydrophilic/hydrophobic balance of the backbone            polymer    -   ii) evaluating the swellability, porosity, and ion binding        interference of the resulting polymer; and    -   iii) selecting a polymer that possesses at least one of the        above features.

In another aspect, the invention provides a method for improving thetherapeutic properties and/or suitability for administration and/orpharmaceutical properties of a polyamine polymer comprising at least oneof the following steps: a) crosslinking said polymer with a crosslinker,such that the average number of connection to the polyamine monomer isbetween about 2.05 and about 6; and/or b) producing said polymer by aprocess wherein the polyamine is initially present in water at a ratioof polyamine:water of from about 3:1 to about 1:3.

C. Monomers

Any suitable monomers and crosslinkers may be used in the polymers ofthe invention. When the polymer binds phosphate or oxalate, the polymerusually comprises a polyamine and a crosslinker. The polyamines includeamine functional monomers such as those described in U.S. Pat. Nos.5,496,545; 5,667,775; 6,509,013; 6,132,706; and 5,968,499; and U.S.patent applications Ser. Nos. 10/806,495 and 10/701,385. These patentsand patent applications are hereby incorporated by reference in theirentirety.

In some embodiments, the invention provides ion-binding polymers thatcontain crosslinked amine moieties. In some of these embodiments, thepolymers are characterized by one or more of the characteristics of lowswelling, high ion binding in vivo, low interference from interferingions, and/or specific porosity. Polymers, including homopolymers andcopolymers, with repeating crosslinked amine units are referred toherein as crosslinked amine polymers. The repeating amine units in thepolymer can be separated by the same or varying lengths of repeatinglinker (or intervening) units. In some embodiments, the polymerscomprise of repeat units of an amine plus intervening linker unit. Inother embodiments, multiple amine units are separated by one or morelinker units.

One monomer useful in the polymers of the invention comprises an amineof formula I

wherein each n, independently, is equal to or greater than 3; m is equalto or greater than 1; and each R₁, independently, is H or optionallysubstituted alkyl or aryl or is linked to a neighboring R₁ to form anoptionally substituted alicyclic, aromatic, or heterocyclic group. Inone embodiment the invention is a crosslinked amine polymer comprisingan amine of Formula I, as described, where the amine is crosslinked witha crosslinking agent.

Preferred amines of formula I include:

In one aspect the invention provides methods of treating an animal,including a human, using the polymers of the invention. One embodimentof this aspect is a method for removing phosphate from thegastrointestinal tract of an animal by administering an effective amountof a crosslinked amine polymer, wherein said polymer comprises an amineof formula I.

A second monomer useful in the polymers of the invention comprises anamine of formula II

wherein p is 1, 2, 3, or 4; each R₁, independently, is H or optionallysubstituted alkyl or aryl or is linked to a neighboring R₁ to form anoptionally substituted alicyclic, aromatic, or heterocyclic group; R₂and R₃, each independently, are H or optionally substituted alkyl oraryl, with the proviso that when p=1, both R₂ and R₃ are not H and whenp=2, 3, or 4, R₂ and R₃ are H, alkyl or —C(R₁)₂—R₄-N(R₁)₂, R₄ beingeither a bond or methylene; in addition, in some of the embodiments, theamines of formula II include amines wherein p is greater than 4. Invarious embodiments, p can be more than 8, more than 12, more than 16,or more than 20. In other embodiments, p can be less than 25, less than20, less than 15, or less than 10. In one embodiment the invention is acrosslinked amine polymer comprising an amine of formula II, asdescribed, where the amine is crosslinked with a crosslinking agent.

Preferred amines of formula II include:

One embodiment of the invention is a method for removing phosphate fromthe gastrointestinal tract of an animal by administering an effectiveamount of a crosslinked amine polymer, wherein said polymer comprises anamine of formula II.

A third monomer useful in the polymers of the invention comprises anamine of formula III

wherein q is 0, 1, or 2; and each R₁, independently, is H or optionallysubstituted alkyl or aryl or is linked to a neighboring R₁ to form anoptionally substituted alicyclic, aromatic, or heterocyclic group. Inone embodiment the invention is a crosslinked amine polymer comprisingan amine of formula III, as described, where the amine is crosslinkedwith a crosslinking agent.

Preferred amines of formula III include:

One embodiment of the invention is a method for removing phosphate fromthe gastrointestinal tract of an animal by administering an effectiveamount of a crosslinked amine polymer, wherein said polymer comprises anamine of formula III.

A fourth monomer useful in the polymers of the invention comprises anamine of formula IV

wherein each n, independently, is equal to or greater than 3; each r,independently, is 0, 1, or 2; and each R₁, independently, is H oroptionally substituted alkyl or aryl or is linked to a neighboring R₁ toform an optionally substituted alicyclic, aromatic, or heterocyclicgroup. In one embodiment the invention is a crosslinked amine polymercomprising an amine of formula IV, as described, where the amine iscrosslinked with a crosslinking agent.

A preferred amine of formula IV includes:

One embodiment of the invention is a method for removing phosphate fromthe gastrointestinal tract of an animal by administering an effectiveamount of a crosslinked amine polymer, wherein said polymer comprises anamine of formula IV.

A fifth monomer useful in the polymers of the invention comprises anamine of formula V

wherein each n, independently, is equal to or greater than 3; each r,independently, is 0, 1, or 2; and each R₁, independently, is H oroptionally substituted alkyl or aryl or is linked to a neighboring R₁ toform an optionally substituted alicyclic, aromatic, or heterocyclicgroup. In one embodiment the invention is a crosslinked amine polymercomprising an amine of formula V, as described, where the amine iscrosslinked with a crosslinking agent.

Preferred amines of formula V include:

One embodiment of the invention is a method for removing phosphate fromthe gastrointestinal tract of an animal by administering an effectiveamount of a crosslinked amine polymer, wherein said polymer comprises anamine of formula V.

A sixth monomer useful in the polymers of the invention comprises anamine of formula VI

wherein each m, independently, is equal to or greater than 3. In oneembodiment the invention is a crosslinked amine polymer comprising anamine of formula VI, as described, where the amine is crosslinked with acrosslinking agent.

One embodiment of the invention is a method for removing phosphate fromthe gastrointestinal tract of an animal by administering an effectiveamount of a crosslinked amine polymer, wherein said polymer comprises anamine of formula VI.

The amines represented by general formulas I-VI can be synthesized bymethods well known in the art. These synthesis techniques includecatalytic conversion from alcohols, reductive amination of carbonylcompounds, Michael additions, and hydrogenation of nitrites (see, forexample, Karsten Eller et al, Ullmann's Encyclopedia of IndustrialChemistry 2002 by Wiley-VCH Verlag GmbH & Co. KGaA). Several small aminemonomers and/or amine plus intervening linker units are alsocommercially available.

In one embodiment, an amine useful in the present invention,tetramethylene tetramine, depicted below, is synthesized by catalytichydrogenation of the commercially available diaminomaleonitrile (DAMN):

Amines that may be used in the present invention are not limited to, butare typically small amines that serve as monomers or parts of monomericunits for the polymerization reactions. In some embodiments, themonomers are low-molecular weight monomers, i.e., monomers of amolecular weight less than 200 g/mol.

In embodiments of the invention, the monomers are non-polymeric, e.g.,non-polymeric amines. As used herein, a “polymer” encompasses a moleculeof high relative molecular mass, the structure of which essentiallycomprises the multiple repetition of units derived, actually orconceptually, from molecules of low relative molecular mass.

Examples of amines that are suitable for synthesis of the polymers ofthe present invention include, but are not limited to, the amines shownin Table 1.

TABLE 1 MW Label Type Structure (g/mol) B-SM-20-TeA Tetramine

316.54 B-SM-22-DA Diamine

61.1 B-SM-23-DA Diamine

88.15 B-SM-24-DA Diamine

74.13 B-SM-25-DA Diamine

88.15 B-SM-26-DA Diamine

129.21 B-SM-27-DA Diamine

114.19 B-SM-28-TA Triamine

196.08 B-SM-29-TA Triamine

125.13 B-SM-31-DA Diamine

184.07 B-SM-32-DA Diamine

136.2

Additional amine monomers that may be used in polymers of the inventioninclude vicinal amine moieties. The polymer may be a homopolymerincluding repeat units of vicinal amines or is a copolymer including oneor more repeat units of vicinal amines and other monomers such asacrylates, methacrylates, acrylamides, methacrylamides, vinyl esters,vinyl amides, olefin, styrenic, etc. The size of the polymer can varybetween, for example, about 500 to about 1,000,000 Daltons.

One vicinal amine monomer useful in the polymers of the invention is themonomer shown in formula VII:

wherein n is zero, one, or greater than 1, each R is independently asuitable chemical group that complements the valency of nitrogen, andeach R′ is independently H, alkyl, or amino.

In another embodiment, the polymer is characterized by a repeating unithaving the formula

or a copolymer thereof, wherein n is zero, one, or greater than 1, eachR is independently a suitable chemical group that complements thevalency of nitrogen, each R′ is independently H, alkyl, or amino, and X⁻is a negatively charged organic or inorganic counterion.

Preferred polymers of formula VIII include:

The polymers of the present invention also include polymerscharacterized by a repeat unit having the formula

wherein n is zero, one, or greater than 1, each R is independently asuitable chemical group that complements the valency of nitrogen, eachR′ is independently H, alkyl, or amino, and X⁻ is a negatively chargedorganic or inorganic counterion.

In one embodiment, the R groups of neighboring nitrogen atoms are linkedto each other to have a structure as depicted in formula X.

wherein Q is a bond, alkyl, alkylamino, alkylcarbonyl, alkenyl, aryl, orheterocyclyl.

In the polymers described herein, n is zero, one, or greater than 1. Inpreferred embodiments, n is 0-5, even more preferably n is zero or 1.

The value of n′ depends on the desired properties of the polymer, thepotential use of the polymer, and the synthesis techniques used.

The pendant nitrogen atom of formulas VII, VIII, IX, and X can be boundto atoms such as C, H, O, S, P and N such that the pendant groups arenitroso, nitro, nitroxide radical, nitrone, nitrene, isocyanate,carbazide, hydrazino, diazo groups, imine, amidine, guanidine,sulfamate, phosphoramidate, and heterocycle.

Examples of suitable R groups include H, halogen, R″, CO₂H, CO₂R″, COR″,C(═N R″)(N R″), CN, CONH₂, CONR″₂, OR″, SO₃R″, Si(R″)₃, and P(O)(OR″)₂.Suitable R″ groups include H, optionally substituted alkyl, acyl,alkylamino, alkenyl, heterocyclyl, and aryl group. Preferred R′ is H,methyl, or amino.

The substituents for R″ groups can be ionic entities with oxygen,nitrogen, phosphorus, or sulfur. Examples of substituents arecarboxylate, sulfonate, sulfamate, sulfone group, phosphonate,phosphazene, phosphoramidate group, quaternary ammonium groups, or aminegroups, e.g., primary and secondary alkyl or aryl amines. Examples ofother suitable substituents include hydroxy, alkoxy, carboxamide,sulfonamide, halogen, alkyl, aryl, hydrazine, guanadine, urea, andcarboxylic acid esters.

Preferred R groups include H and the following groups:

The negatively charged counterions, X⁻, can be organic ions, inorganicions, or a combination thereof. The inorganic ions suitable for use inthis invention include halide (especially chloride), carbonate,bicarbonate, sulfate, bisulfate, hydroxide, nitrate, persulfate andsulfite. Suitable organic ions include acetate, ascorbate, benzoate,citrate, dihydrogen citrate, hydrogen citrate, oxalate, succinate,tartrate, taurocholate, glycocholate, and cholate. Preferred X⁻ ischloride or carbonate.

In a preferred embodiment, the counterion does not have a detrimentalside effect to the patient and is selected to have a therapeutic ornutritional benefit to the patient.

Another monomer of use in the polymers of the invention is formula XIshown below,

wherein R′″ is H or CH₃, and R has the same meaning as above. Preferredstructures of formula XI are when R═H.

In one embodiment, the polymer is a copolymer with one of the repeatunits being a monomer as described herein.

The copolymers of the present invention can be alternative or randomcopolymers. Generally, monomers that may be co-polymerized with theamine precursors include one or more monomers selected from the groupconsisting of styrene, substituted styrene, alkyl acrylate, substitutedalkyl acrylate, alkyl methacrylate, substituted alkyl methacrylate,acrylonitrile, methacrylonitrile, acrylamide, methacrylamide,N-alkylacrylamide, N-alkylmethacrylamide, N,N-dialkylacrylamide,N,N-dialkylmethacrylamide, isoprene, butadiene, ethylene, vinyl acetate,N-vinyl amide, maleic acid derivatives, vinyl ether, allyle, methallylmonomers and combinations thereof. Functionalized versions of thesemonomers may also be used. Specific monomers or comonomers that may beused in this invention include, but are not limited to, methylmethacrylate, ethyl methacrylate, propyl methacrylate (all isomers),butyl methacrylate (all isomers), 2-ethylhexyl methacrylate, isobornylmethacrylate, methacrylic acid, benzyl methacrylate, phenylmethacrylate, methacrylonitrile, α-methylstyrene, methyl acrylate, ethylacrylate, propyl acrylate (all isomers), butyl acrylate (all isomers),2-ethylhexyl acrylate, isobornyl acrylate, acrylic acid, benzylacrylate, phenyl acrylate, acrylonitrile, styrene, glycidylmethacrylate, 2-hydroxyethyl methacrylate, hydroxypropyl methacrylate(all isomers), hydroxybutyl methacrylate (all isomers),N,N-dimethylaminoethyl methacrylate, N,N-diethylaminoethyl methacrylate,triethyleneglycol methacrylate, itaconic anhydride, itaconic acid,glycidyl acrylate, 2-hydroxyethyl acrylate, hydroxypropyl acrylate (allisomers), hydroxybutyl acrylate (all isomers), N,N-dimethylaminoethylacrylate, N,N-diethylaminoethyl acrylate, triethyleneglycol acrylate,methacrylamide, N-methylacrylamide, N,N-dimethylacrylamide,N-tert-butylmethacrylamide, N-n-butylmethacrylamide,N-methylolmethacrylamide, N-ethylolmethacrylamide,N-tert-butylacrylamide, N-n-butylacrylamide, N-methylolacrylamide,N-ethylolacrylamide, 4-acryloylmorpholine, vinyl benzoic acid (allisomers), diethylaminostyrene (all isomers), α-methylvinyl benzoic acid(all isomers), diethylamino α-methylstyrene (all isomers),p-vinylbenzene sulfonic acid, p-vinylbenzene sulfonic sodium salt,trimethoxysilylpropyl methacrylate, triethoxysilylpropyl methacrylate,tributoxysilylpropyl methacrylate, dimethoxymethylsilylpropylmethacrylate, diethoxymethylsilylpropyl methacrylate,dibutoxymethylsilylpropyl methacrylate, diisopropoxymethylsilylpropylmethacrylate, dimethoxysilylpropyl methacrylate, diethoxysilylpropylmethacrylate, dibutoxysilylpropyl methacrylate, diisopropoxysilylpropylmethacrylate, trimethoxysilylpropyl acrylate, triethoxysilylpropylacrylate, tributoxysilylpropyl acrylate, dimethoxymethylsilylpropylacrylate, diethoxymethylsilylpropyl acrylate, dibutoxymethylsilylpropylacrylate, diisopropoxymethylsilylpropyl acrylate, dimethoxysilylpropylacrylate, diethoxysilylpropyl acrylate, dibutoxysilylpropyl acrylate,diisopropoxysilylpropyl acrylate, maleic anhydride, N-phenylmaleimide,N-butylmaleimide, N-vinylformamide, N-vinyl acetamide, allylamine,methallylamine, allylalcohol, methyl-vinylether, ethylvinylether,butylvinyltether, butadiene, isoprene, chloroprene, ethylene, vinylacetate and combinations thereof. The preferred monomers or comonomersare acrylamide, dimethylacrylamide, N-vinyl formamide, N-vinylacetamide,vinyl acetate, methyl acrylate, and butyl acrylate.

Further monomers that may be used in the polymer of the inventioninclude:

where each R, independently, is H or a substituted or unsubstitutedalkyl, such as a lower alkyl (e.g., having between 1 and 5 carbon atoms,inclusive), alkylamino (e.g., having between 1 and 5 carbons atoms,inclusive, such as ethylamino) or aryl (e.g., phenyl) group;

where each R, independently, is H or a substituted or unsubstitutedalkyl (e.g., having between 1 and 5 carbon atoms, inclusive), alkylamino(e.g., having between 1 and 5 carbons atoms, inclusive, such asethylamino) or aryl (e.g., phenyl) group, and each X⁻ is an exchangeablenegatively charged counterion.

Another suitable monomer is a structure of the formula

where R is H or a substituted or unsubstituted alkyl (e.g., havingbetween 1 and 5 carbon atoms, inclusive), alkylamino (e.g., havingbetween 1 and 5 carbons atoms, inclusive, such as ethylamino) or arylgroup (e.g., phenyl).

Another suitable monomer is a structure of the formula

where each R₁ and R₂, independently, is H or a substituted orunsubstituted alkyl (e.g., having between 1 and 5 carbon atoms,inclusive), and alkylamino (e.g., having between 1 and 5 carbons atoms,inclusive, such as ethylamino) or aryl group (e.g., phenyl), and each X⁻is an exchangeable negatively charged counterion. In one embodiment, atleast one of the R groups is a hydrogen atom.

Another suitable monomer is a structure of the formula

where each R₁ and R₂, independently, is H, a substituted orunsubstituted alkyl group containing 1 to 20 carbon atoms, an alkylaminogroup (e.g., having between 1 and 5 carbons atoms, inclusive, such asethylamino), or an aryl group containing 6 to 12 atoms (e.g., phenyl).

Another suitable monomer is a structure of the formula

where each R₁ and R₂ and R₃, independently, is H, a substituted orunsubstituted alkyl group containing 1 to 20 carbon atoms, an alkylaminogroup (e.g., having between 1 and 5 carbons atoms, inclusive, such asethylamino), or an aryl group containing 6 to 12 atoms (e.g., phenyl),and each X⁻ is an exchangeable negatively charged counterion.

In each case for these monomers, the R groups can carry one or moresubstituents. Suitable substituents include therapeutic anionic groups,e.g., quaternary ammonium groups, or amine groups, e.g., primary andsecondary alkyl or aryl amines. Examples of other suitable substituentsinclude hydroxy, alkoxy, carboxamide, sulfonamide, halogen, alkyl, aryl,hydrazine, guanadine, urea, and carboxylic acid esters, for example.

The negatively charged counterions, X⁻, can be organic ions, inorganicions, or a combination thereof. The inorganic ions suitable for use inthis invention include halide (especially chloride), carbonate,bicarbonate, sulfate, bisulfate, hydroxide, nitrate, persulfate andsulfite. Suitable organic ions include acetate, ascorbate, benzoate,citrate, dihydrogen citrate, hydrogen citrate, oxalate, succinate,tartrate, taurocholate, glycocholate, and cholate.

Polymers containing guanidino groups are also useful as compositionsthat may be produced by the processes described herein to have thedesired properties, and that bind anions such as phosphate and oxalate.Such polymers are described in U.S. Pat. Nos. 6,132,706; and 5,968,499,which are hereby incorporated by reference in their entirety. Briefly,guanidino groups are attached to a polymeric structure. The nature ofthe polymeric backbone is not of primary importance as the bindingeffect is due to the guanidino groups. Preferred polymers in whichcrosslinking and other factors may be controlled include polymers havinga polyethylene backbone crosslinked with divinyl benzene. Polymershaving an inorganic backbone, for example the polyphosphazene polymers,may also be used. The polymers may be copolymers derived from two ormore different types of monomer. Further examples of useful polymers arecarbohydrate polymers including cellulose and agarose. The guanidinogroups are attached to the polymer backbone by means of chemical bondingthrough the terminal NH group of the guanidino group (NH₂—C(═NH)—NH—).The chemical bonding of the guanidino groups to the polymer backbone maybe directly or via some form of grouping acting as a “spacer” throughwhich it is attached to the polymer backbone. Various forms ofattachment may be used, preferred forms varying according to the basictype of polymer. For example, alkylene groups of 1-4 carbon atoms, amidegroups, ether groups or a combination thereof may be used. The preferredmode of attachment of guanidino groups to the polymer backbone willobviously depend upon the nature of the backbone but for simplicitydirect bonding between atoms of the backbone and the NH group of theguanidino group is preferred where possible.

Methods of preparing the guanidino-containing polymers will be apparentto a person skilled in the art but for example, they may be preparedfollowing the teachings of Schnaar, R. L. and Lee, Y. C., 1975,Biochemistry 14, 1535-1541, hereby incorporated by reference in itsentirety, who describes a method for linking biologically active ligandsto a polymer matrix, or the polymers may also conveniently be preparedthrough the reaction with a polymer containing amino groups attached tothe polymer backbone of (a) 3,5-dimethylpyrazole-1-carboxamidinenitrate, (b) S-methylthiouronium sulphate or (c) O-methylpseudoureahydrogen sulphate.

Preferred monomers of the invention are amines. Most preferred monomersfor use in the polymers of the invention include allylamine, vinylamine,ethyleneimine, methylene 1,3 diamino propane, and N,N,N′,N′-tetrakis(3-aminopropyl) 1,4 diaminobutane, 1,2,3,4 tetraminobutane, formula 1and formula 2, wherein formula 1 and formula 2 are the followingstructures:

In some embodiments, polymers of the invention are composed of one ormore amine monomers and one or more crosslinkers where the polymer isproduced by a process in which the amine is present in solvent beforecrosslinking at a ratio of amine:solvent of from about 3:1 to about 1:3and the total content crosslinkers added to the reaction mix is suchthat the average number of connections to the amine monomers is betweenabout 2.05 and about 6, or between about 2.2 and about 4.5. In someembodiments, polymers of the invention are a phosphate-binding polymercomposed of one or more amine monomers and one or more crosslinkerswhere the polymer is produced by a process wherein the total contentcrosslinkers added to the reaction mix is such that the average numberof connections to the amine monomers is between 2.2 and 4.5. Inpreferred embodiments, the amine monomer is selected from the groupconsisting of 1,3 diamino propane, and N,N,N′,N′-tetrakis(3-aminopropyl) 1,4 diaminobutane, and wherein the crosslinker isselected from the group consisting of 1,3 dichloropropane andepichlorohydrin. In some embodiments, polymers of the invention arecomposed of one or more amine monomers and one or more crosslinkers,wherein the amine monomers are not polyallylamine monomers and/or thecrosslinkers are not epichlorhydrin.

In some embodiments, e.g., in phosphate-binding polymers, it isdesirable to keep the chloride to amine ratio of the final polymer belowcertain levels. In some embodiments, this is about 0 to about 35 mol %,preferably about 0 to about 15 mol %. Monomers may be selected accordingto this criterion.

D. Crosslinkers

The crosslinker include those described in U.S. Pat. Nos. 5,496,545;5,667,775; 6,509,013; 6,132,706; and 5,968,499; and U.S. patentapplications Ser. Nos. 10/806,495 and 10/701,385.

Crosslinking agents are typically compounds having at least twofunctional groups that are selected from a halogen group, carbonylgroup, epoxy group, ester group, acid anhydride group, acid halidegroup, isocyanate group, vinyl group, and chloroformate group. Thecrosslinking agent may be attached to the carbon backbone or to thependant nitrogen of the amine polymer. Examples of crosslinkers that aresuitable for synthesis of the polymers of the present invention include,but are not limited to, the crosslinkers shown in Table 2.

TABLE 2 Label Structure Mw X-EP-1

92.52 X-EP-2

174.19 X-EP-3

X-EP-4

302.37 X-EP-5

297.27 X-EP-6

277.32 X-EP-7

86.09 X-EP-8

202.25 X-Cl-1

184.41 X-Cl-2

175.06 X-Cl-3

112.99 X-Cl-4

178.49 X-Cl-5

240.99 X-Cl-6

127.01 X-AC-1

203.02 X-AC-2

203.02 X-AC-3

265.48 X-AC-4

154.98 X-AH-1

198.13 X-AH-2

X-AH-3

112.08 X-Mc-1

168.2 X-Mc-2

118.16 X-Mc-3

249.27 X-IC-1

168.19 X-IC-2

174.16 X-IC-3

188.18 X-IC-4

222.28 X-ME-1

86.09 X-ME-2

158.16 X-ME-3

146.14 X-ME-4

194.19 X-ME-5

234.2 X-ME-6

252.22 X-ME-7

194.19 X-ME-8

178.14 X-ME-9

108.53

Examples of suitable crosslinking agents are diacrylates anddimethacrylates (e.g., ethylene glycol diacrylate, propylene glycoldiacrylate, butylene glycol diacrylate, ethylene glycol dimethacrylate,propylene glycol dimethacrylate, butylene glycol dimethacrylate,polyethyleneglycol dimethacrylate, polyethyleneglycol diacrylate),methylene bisacrylamide, methylene bismethacrylamide, ethylenebisacrylamide, ethylenebismethacrylamide, ethylidene bisacrylamide,divinyl benzene, bisphenol A dimethacrylate, bisphenol A diacrylate,diepoxides, dihalides, diisocyanates, diacyl chlorides, dianhydrides,and dimethyl esters.

Examples of preferred crosslinking agents include epichlorohydrin, 1,4butanedioldiglycidyl ether, 1,2 ethanedioldiglycidyl ether,1,3-dichloropropane, 1,2-dichloroethane, 1,3-dibromopropane,1,2-dibromoethane, succinyl dichloride, dimethylsuccinate, toluenediisocyanate, acryloyl chloride, methyl acrylate, ethylenebisacrylamide, and pyromellitic dianhydride.

E. Polymerization

Polymerization can be achieved by methods known in the art, examples ofwhich are illustrated in detail in the Examples disclosed herein. Asdescribed above, polymerization conditions may be manipulated in orderto produce polymers with the desired characteristics.

The crosslinking reaction is carried out either in bulk solution (i.e.using the neat amine and neat crosslinker compounds) or in dispersedmedia. The crosslinking reaction leading to gel formation can beperformed using a variety of processes; these processes fall into twocategories:

i) homogeneous processes where the amine functional precursor (smallmolecule amine or high molecular weight polyamine) is soluble in thecontinuous phase, and where the gel, obtained by a crosslinkingreaction, is recovered as a bulk gel or gel slurry in said continuousphase. Bulk gel process describes situation where the entirety of thesolvent is trapped in the gel network forming a mass that is thencomminuted in smaller particles using extrusion, grinding and relatedmethods. When a bulk process is used, solvents are selected so that theyco-dissolve the reactants and do not interfere with the aminecrosslinking reaction. Suitable solvents include water, low boilingalcohols (methanol, ethanol, butanol), dimethylformamide,dimethylsulfoxide, acetone, methylethylketone, and the like. A gelslurry is typically obtained where the reaction medium viscosity is thelow range and the shear rate high so that pieces of gel are producedthat stay in suspension in a slurry form.

ii) heterogeneous processes, where the amine functional precursor (smallmolecule amine or high molecular weight polyamine) is made insoluble inthe continuous phase so as to form dispersed droplets or particles,which then undergo a crosslinking reaction, forming bead orirregularly-shaped particles kept in suspension in said continuousphase.

Homogeneous processes can be impractical for crosslinked material withlimited swelling ratios such as those contemplated in this invention:the level of crosslinking typical for the desired range of swellingratio and pore size distribution usually induces very short gel time andhigh local viscosity, both of which are impractical in large scalemanufacturing.

A preferred mode of synthesis for the present invention is to useheterogeneous processes. Such processes are also referred to aspolymerization in dispersed media and include inverse suspension, directsuspension, precipitation polymerization, emulsion polymerization andmicroemulsion polymerization, reaction in aerosols, and the like. Thecontinuous phase can be selected from a polar solvents such as toluene,benzene, hydrocarbon, halogenated solvents, supercritical carbondioxide, and the like. With a direct suspension or emulsion process,water can be used, although salt brines are also useful to “salt out”the amine and crosslinker reagents in a droplet separate phase, asdescribed in U.S. Pat. No. 5,414,068. The monomer precursors can bedispersed either neat or as a solution in the continuous phase. Theamine and crosslinker are preferably introduced in two separate steps,wherein the amine is first dispersed as droplets, and subsequently thecrosslinker is added to the reaction medium and migrates to thedispersed phase. The crosslinking reaction occurs within the dropletphase without causing any significant increase in viscosity in thedispersion. This has the advantage of dissipating the heat generated bythe exothermic reaction while insuring good gel homogeneity within thebeads. A preferred mode of synthesis comprises the steps of:

-   -   i) solubilizing the amine monomer or amine polymer in water    -   ii) neutralizing a fraction of the amine with an acid, such as        HCl,    -   iii) dispersing said amine solution in a water immiscible        solvent to form an emulsion    -   iv) adding the crosslinker to the emulsion in a staged addition    -   v) allowing the crosslinking reaction to proceed to completion    -   vi) removing the water by distillation    -   vii) isolating the beads by filtration    -   viii) washing and drying

In this process the polymer particles are obtained as spherical beads,whose diameter is preferably controlled in the 5 to 500 microns range,preferably 25 to 250 microns. In some of these embodiments the beadshave a mean diameter of less than 40 microns.

Thus, in one, aspect, the invention provides a method of making ananion-binding polymer that binds a target anion, comprising combining anamine monomer with a crosslinker by a heterogeneous process, wherein thephosphate-binding polymer is characterized by at least two of thefollowing features: a) a swelling ratio of less than about 5, or lessthan about 4.5, or less than about 4, or less than about 3; b) less thanabout 20% of the weight of the polymer accessible to non-interactingsolutes of molecular weight greater than about twice the MW of thetarget anion, wherein said percentage is measured in a physiologicalmedium, and c) an ion-binding interference for the target anion lowerthan about 60% when measured in a gastrointestinal simulant, relative toa non-interfering buffer. In some embodiments the amine monomer is apolyallylamine. In some embodiments the crosslinker is epichlorohydrin.

In another aspect the invention provides an anion-binding polymer thatbinds a target ion, wherein the polymer is produced by a processcomprising crosslinking a polyallylamine by a heterogeneous process, andwherein said polymer is characterized by at least two of the followingfeatures: a) a swelling ratio of less than about 5, or less than about4.5, or less than about 4, or less than about 3; b) less than about 20%of the weight of the polymer accessible to non-interacting solutes ofmolecular weight greater than about twice the MW of the target anion,wherein said percentage is measured in a physiological medium, and c) anion-binding interference for the target anion lower than about 60% whenmeasured in a gastrointestinal simulant, relative to a non-interferingbuffer. In one embodiment, the polyallyamine is crosslinked byepichlorohydrin.

As discussed above, the crosslinker to amine mole ratios control theextent of gel material formed as well as its crosslinking density. Toolow a ratio may lead to incomplete crosslinking and formation of solubleoligomers, while too high a ratio may produce an extremely tight networkwith little binding properties. The amine component can be either one ora combination of several amines, and the same applies to the crosslinkercomponent. Optimization may be required for any new combination ofamines and crosslinkers, since the functionality of either can influencethe extent of gel formation and swelling characteristics. In someembodiments, e.g., embodiments of low molecular weight monomerscrosslinked by crosslinkers with an Fb of 2, crosslinker to amine molarratios (B/A) comprise between about 0.2 to about 10, preferably about0.5 to about 5, and most preferably about 0.5 to about 2. These ratiosmay be adjusted, based on whether the amine monomer is a high molecularweight or low molecular weight monomer, and/or the Fb number of thecrosslinker (see discussion and table above).

In some cases the polymers are crosslinked after polymerization. Onemethod of obtaining such crosslinking involves reaction of the polymerwith difunctional crosslinkers, such as epichlorohydrin, succinyldichloride, the diglycidyl ether of bisphenol A, pyromelliticdianhydride, toluene diisocyanate, and ethylenediamine. A typicalexample is the reaction of poly(ethyleneimine) with epichlorohydrin. Inthis example the epichlorohydrin (1 to 100 parts) is added to a solutioncontaining polyethyleneimine (100 parts) and heated to promote reaction.A typical example is the reaction of polyvicinalamine withepichlorohydrin. In this example the epichlorohydrin (1 to 200 parts) isadded to a solution containing polyvicinalamine (100 parts) and heatedto promote reaction. Other methods of inducing crosslinking on alreadypolymerized materials include, but are not limited to, exposure toionizing radiation, ultraviolet radiation, electron beams, radicals, andpyrolysis.

The crosslinking reaction is run in a batch or semi continuous mode. Inthe latter mode, either the amine or the crosslinker is added as theinitial charge and the co-reactant is then metered up for a given periodof time. In one embodiment, a soluble prepolymer is first prepared byadding the entire amine monomer component and then adding continuously afraction of the crosslinker, forming a syrup. The syrup is thenemulsified as droplets in an oil continuous phase and the remainingfraction of crosslinker is added to form crosslinked beads. When thecrosslinker is an alkylhalide compound, a base can be used to scavengethe acid formed during the reaction. Inorganic or organic bases aresuitable. NaOH is preferred. The base to crosslinker ratio is preferablybetween about 0.5 to about 2.

In some embodiments the polymers are subject to post-amination(post-reaction with 3-chloropropylamine). In this embodiment, a firstreaction between an amine monomer and a crosslinker is carried out toform a gel, then the gel is post reacted with an aminoalkylhalide, wherethe amine alkyl groups are chemically attached to the gel through halidesubstitution by the amine functional gels.

All of the polymers described herein carl be further cross-linked andimpTinted with anion, e.g., phosphate. In one embodiment the targetanion (e.g., phosphate or oxalate) is present dining the polymerizationand is then washed out when the crosslinking reaction is completed. Ihemethod is referred to as “imprinting” and tends to increase the chemicalaffinity of the gel towards the anion solute by creating “molded”pockets within the gel that have high binding recognition for a givenanion Examples of phosphate imprinted gels are described in e.g.Fujiwara et al, Analytical Sciences April 2000, vol. 16, 407, and in ACSsymposium series 703, “Molecular and Ionic Recognition with ImprintedPolymers, Bartsch RA and Maeda M. Editors, 1998, Chap. 22, 315.Typically the anion is present at a mole ratio to amine (expressed asnitrogen atom) of between about 10% to about 100%, more preferably about10% to about 60%, most preferably about 30% to about 50% Mostpreferably, the anion is introduced in the acid fbrm (e.g., phosphoricacid, oxalic acid) and the amine as the free base, so as to form, insitu, the ammonium/anion salt. Crosslinking is then carried out asdescribed earlier by using the proper amount crosslinker to amine molarratio in order to obtain the desired gel features in terms of swellingratio, critical permeation volume, and binding interference. The gelimmediately formed after crosslinking is then thoroughly washed ineither highly acidic (e.g., pH<2) or highly basic (e.g., pH>12) mediumto remove the imprinted anion, then frither washed with neutral medium.All parameters being equal (e.g., amine to crosslinker ratio, monomer tosolvent ratio) the imprinting method here described usually increasesthe binding capacity by a factor of 1.1, 1.3, or even 1.5.

III. Pharmaceutical Compositions

In one aspect the invention provides pharmaceutical compositions. In oneembodiment, the pharmaceutical compositions are chewable tablets. Inanother embodiment, the pharmaceutical compositions are liquidformulations.

The pharmaceutical compositions of the present invention includecompositions wherein the polymers of the invention, e.g., crosslinkedamine polymers, are present in an effective amount, i.e., in an amounteffective to achieve therapeutic and/or prophylactic benefit. The actualamount effective for a particular application will depend on the patient(e.g. age, weight) the condition being treated; and the route ofadministration. Determination of an effective amount is well within thecapabilities of those skilled in the art, especially in light of thedisclosure herein.

The effective amount for use in humans can be determined from animalmodels. For example, a dose for humans can be formulated to achievecirculating and/or gastrointestinal concentrations that have been foundto be effective in animals.

The pharmaceutical compositions comprise the polymer, e.g., crosslinkedamine polymers, one or more pharmaceutically acceptable carriers,diluents or excipients, and optionally additional therapeutic agents.

Pharmaceutical compositions for use in accordance with the presentinvention may be formulated in conventional manner using one or morephysiologically acceptable carriers comprising excipients andauxiliaries which facilitate processing of the active compounds intopreparations which can be used pharmaceutically. Proper formulation isdependent upon the route of administration chosen. Suitable techniquesfor preparing pharmaceutical compositions of the amines are well knownin the art, e.g., Gennaro A R (ed), Remington's Pharmaceutical Sciences,20th Edition, Lippincott, Williams and Wilkins, Baltimore Md. (2001),which is hereby incorporated in its entirety.

The present pharmaceutical compositions are generally prepared by knownprocedures using well known and readily available ingredients. In makingthe compositions of the present invention, the ion-binding polymer,e.g., a phosphate-binding polymer, may be present alone, may be admixedwith a carrier, diluted by a carrier, or enclosed within a carrier whichmay be in the form of a capsule, sachet, paper or other container. Whenthe carrier serves as a diluent, it may be a solid, semi-solid or liquidmaterial which acts as a vehicle, excipient or medium for the polymer.Thus, the compositions can be in the form of tablets, pills, powders,lozenges, sachets, cachets, elixirs, suspensions, syrups, aerosols, (asa solid or in a liquid medium), soft or hard gelatin capsules, sterilepackaged powders, and the like. Preferred formulations are chewabletablets and liquid formulations. Examples of carriers, excipients, anddiluents that may be used in these formulations as well as others,include foods, drinks, lactose, dextrose, sucrose, sorbitol, mannitol,starches, gum acacia, alginates, tragacanth, gelatin, calcium silicate,microcrystalline cellulose, polyvinylpyrrolidone, cellulose, methylcellulose, methylhydroxybenzoates, propylhydroxybenzoates,propylhydroxybenzoates, and talc.

In another aspect of the invention, the anion (e.g., phosphate) bindingpolymer is formulated as the free amine, free of counter-ions. Shortterm and long term studies have demonstrated that maintenancehemodialysis patients treated with Renagel (polyallylaminehydrochloride) have significantly lower serum bicarbonate levels thanpatients treated with calcium-containing phosphate binders (i.e.containing no chloride). It has been shown (Brezina B. et al, KidneyInternational, vo. 66, suppl. 90 (2004), 39-45) that SEVELAMERhydrochloride (Trade name of Renagel active pharmaceutical ingredient)induces an acid load that causes acidosis. Acidosis can have seriousside effects for that category of patients. In another embodiment, theamine crosslinked polymer is a polyamine polymer where the chloridecontent of the polymer is less than about 40 mol % of the amine groupcontent, more preferably less than about 20 mol % of the amine groupcontent, and even more preferably less than about 5% of the amine groupcontent. Most preferably, the polymer is substantially chloride-free.

A. Chewable Tablets

In some embodiments the polymers of the invention are provided aspharmaceutical compositions in the form of chewable tablets.

Patient compliance is recognized today as one of the main limitingfactors for patients to adhere to recommendations in treating ionimbalance disorders, such as hyperphosphatemia. For example, in thetreatment of hyperphosphatemia using current phosphate-binding polymers,such as RENAGEL recent surveys imply that patients have to take onaverage nine to ten 800 mg pills per day with 25% of the patientpopulation taking even higher daily doses of twelve to fifteen pills.Renagel takes the form of swallowable tablets and is administered withamounts of fluid necessary to ingest the tablets, adding to the burdenof ESRD patients who are under fluid restriction. Low patient compliancedue to large daily doses stands out as a factor that clearly impactsacceptance of this class of drugs.

More easy-to-take pharmaceutical formulation would be desirable. Thoughdrug delivery by chewable tablet would be highly advantageous in manycases, usage has been limited as formulators have encountereddifficulties in achieving satisfactory sensory characteristics. Whenchewing a tablet the following sensory parameters are important:grittiness, tooth packing, chalkiness, mouthfeel, and overallpalatability.

Current chewable tablets are mostly used in areas where significantamounts of active ingredients need to be administered and include overthe counter products such as vitamins, antacids, laxatives and painmedications. Prescription chewable products include prenatal vitaminsand chewable antibiotic and antiviral products that require large dosesto be orally administered. Although often large, the geometry needs tobe optimized to facilitate ease of chewing and “hardness” appropriatefor mastication. Round bevel-edged shapes are common withheight/diameter ratios around 0.3 to 0.4.

In addition to the active ingredient, the following types of excipientsare commonly used: a sweetening agent to provide the necessarypalatability, plus a binder where the former is inadequate in providingsufficient tablet hardness; a lubricant to minimize frictional effectsat the die wall and facilitate tablet ejection; and, in someformulations a small amount of a disintegrant is added to facilitatemastication. In general excipient levels in currently-available chewabletablets are on the order of 3-5 fold of active ingredient(s) whereassweetening agents make up the bulk of the inactive ingredients.

An important consideration in designing a chewable tablet containing anion-binding polymer is the swelling ratio of the polymer. Because theinvention provides polymers that are low-swelling, they may be used inchewable formulations without the unpleasant and sometimes dangerousside effects that accompany chewable tablets of higher-swellingpolymers. One example of a high swelling material causing difficultiesduring oral administration potentially resulting in choking and blockageof the esophagus is Psyllium. Psyllium comes from the crushed seeds ofthe Plantago ovata plant, an herb native to parts of Asia, Mediterraneanregions of Europe, and North Africa and is commonly used as a laxativein the US. Psyllium typically swells 35-50 times and has to be takenwith plenty of fluids. Insufficient fluid uptake upon administration maycause the fiber to swell and result in choking or even rupture of theesophagus. Psyllium is contraindicated in patients that have dysphagiaand/or a narrow esophagus.

The present invention provides chewable tablets that contain a polymeror polymers of the invention and one or more pharmaceutical excipientssuitable for formulation of a chewable tablet. The polymer used inchewable tablets of the invention preferably has a swelling ratio whiletransiting the oral cavity and in the esophagus of less than about 5,preferably less than about 4, more preferably less than about 3, morepreferably less than 2.5, and most preferably less than about 2. In someembodiments the polymer is an anion-binding polymer such as a phosphate-or oxalate binding polymer; in a preferred embodiment, the polymer is aphosphate-binding polymer. The tablet comprising the polymer, combinedwith suitable excipients, provides acceptable organoleptic propertiessuch as mouthfeel, taste, and tooth packing, and at the same time doesnot pose a risk to obstruct the esophagus after chewing and contact withsaliva.

In some aspects of the invention, the polymer(s) provide mechanical andthermal properties that are usually performed by excipients, thusdecreasing the amount of such excipients required for the formulation.In some embodiments the active ingredient (e.g., polymer, preferably ananion-binding polymer) constitutes over about 30%, more preferably overabout 40%, even more preferably over about 50%, and most preferably morethan about 60% by weight of the chewable tablet, the remaindercomprising suitable excipient(s). In some embodiments the polymer, e.g.,an anion-binding polymer, comprises about 0.6 gm to about 2.0 gm of thetotal weight of the tablet, preferably about 0.8 gm to about 1.6 gm. Insome embodiment the polymer, e.g., an anion-binding polymer, comprisesmore than about 0.8 gm of the tablet, preferably more than about 1.2 gmof the tablet, and most preferably more than about 1.6 gm of the tablet.The polymer is produced to have appropriate strength/friability andparticle size to provide the same qualities for which excipients areoften used, e.g., proper hardness, good mouth feel, compressibility, andthe like. Particle size for polymers used in chewable tablets of theinvention is less than about 80, 70, 60, 50, 40, 30, or 20 microns meandiameter. In preferred embodiments, the particle size is less than about80, more preferably less than about 60, and most preferably less thanabout 40 microns.

Pharmaceutical excipients useful in the chewable tablets of theinvention include a binder, such as microcrystalline cellulose,colloidal silica and combinations thereof (Prosolv 90), carbopol,providone and xanthan gum; a flavoring agent, such as sucrose, mannitol,xylitol, maltodextrin, fructose, or sorbitol; a lubricant, such asmagnesium stearate, stearic acid, sodium stearyl fumurate and vegetablebased fatty acids; and, optionally, a disintegrant, such ascroscarmellose sodium, gellan gum, low-substituted hydroxypropyl etherof cellulose, sodium starch glycolate. Other additives may includeplasticizers, pigments, talc, and the like. Such additives and othersuitable ingredients are well-known in the art; see, e.g., Gennaro A R(ed), Remington's Pharmaceutical Sciences, 20th Edition.

In some embodiments the invention provides a pharmaceutical compositionformulated as a chewable tablet, comprising a phosphate-binding polymerand a suitable excipient. In some embodiments the invention provides apharmaceutical composition formulated as a chewable tablet, comprising aphosphate-binding polymer, a filler, and a lubricant. In someembodiments the invention provides a pharmaceutical compositionformulated as a chewable tablet, comprising a phosphate-binding polymer,a filler, and a lubricant, wherein the filler is chosen from the groupconsisting of sucrose, mannitol, xylitol, maltodextrin, fructose, andsorbitol, and wherein the lubricant is a magnesium fatty acid salt, suchas magnesium stearate.

The tablet may be of any size and shape compatible with chewability andmouth disintegration, preferably of a cylindrical shape, with a diameterof about 10 mm to about 40 mm and a height of about 2 mm to about 10 mm,most preferably a diameter of about 22 mm and a height of about 6 mm.

In one embodiment the polymer has a transition temperature greater thanabout 30° C., preferably greater than about 50° C.

In another embodiment, the polymer is pre-formulated with a high Tg/highmelting point low molecular weight excipient such as mannitol, sorbose,sucrose in order to form a solid solution wherein the polymer and theexcipient are intimately mixed. Method of mixing such as extrusion,spray-drying, chill drying, lyophilization, or wet granulation areuseful. Indication of the level of mixing is given by known physicalmethods such as differential scanning calorimetry or dynamic mechanicalanalysis.

Methods of making chewable tablets containing pharmaceuticalingredients, including polymers, are known in the art. See, e.g.,European Patent Application No. EP373852A2 and U.S. Pat. No. 6,475,510,and Remington's Pharmaceutical Sciences, which are hereby incorporatedby reference in their entirety.

B. Liquid Formulations

In some embodiments the polymers of the invention are provided aspharmaceutical compositions in the form of liquid formulations. In someembodiments the pharmaceutical composition contains an ion-bindingpolymer dispersed in a suitable liquid excipient. Suitable liquidexcipients are known in the art; see, e.g., Remington's PharmaceuticalSciences.

IV. Methods of Treatment

In another aspect, the invention provides methods of treatment of ionimbalance disorders. The term “ion imbalance disorders” as used hereinrefers to conditions in which the level of an ion present in the body isabnormal. In one embodiment, the invention provides methods of treatinga phosphate imbalance disorder. The term “phosphate imbalance disorder”as used herein refers to conditions in which the level of phosphoruspresent in the body is abnormal. One example of a phosphate imbalancedisorder includes hyperphosphatemia. The term “hyperphosphatemia” asused herein refers to a condition in which the element phosphorus ispresent in the body at an elevated level. Typically, a patient is oftendiagnosed with hyperphosphatemia if the blood phosphate level is, forexample, above about 4.5 milligrams per deciliter of blood and/orglomerular filtration rate is reduced to, for example, more than about20%.

Thus, for example, the invention provides methods of removing an anionfrom an animal by administering an effective amount of a polymer of theinvention to the animal. In some embodiments, the polymer is ananion-binding polymer where the polymer binds a target anion (e.g.,phosphate or oxalate), and where the polymer is characterized by atleast two of the following features: a) a swelling ratio of less thanabout 5; b) a gel pore volume distribution measured in a physiologicalmedium characterized by a fraction of said pore volume accessible tonon-interacting solutes, of molecular weight greater than about twicethe MW of the target anion, of less than about 20% of the weight of thegel; and c) an ion-binding interference for the target anion lower thanabout 60% when measured in a gastrointestinal simulant, relative to anon-interfering buffer. In some embodiments, the target anion of thepolymer is phosphate; in some embodiments the phosphate is removed fromthe gastrointestinal tract; in some embodiments the method ofadministration is oral. In some embodiments, the animal is afflictedwith at least one disease selected from the group consisting ofhyperphosphatemia, hypocalcemia, hyperthyroidism, depressed renalsynthesis of calcitriol, tetany due to hypocalcemia, renalinsufficiency, ectopic calcification in soft tissues, and ESRD. In someembodiments, the animal is a human. It will be appreciated that anypolymer described herein may be useful in binding an anion in an animaland/or in treating conditions caused by an ion imbalance in an animal.In preferred embodiments, the polymer is a phosphate-binding polymerwhere the polymer is characterized by at least one of the followingfeatures: a) a swelling ratio of less than about 5, preferably less thanabout 2.5; b) a gel pore volume distribution measured in a physiologicalmedium characterized by a fraction of said pore volume accessible tonon-interacting solutes, of molecular weight greater than about 200, ofless than about 20% of the weight of the gel; and c) an ion-bindinginterference for phosphate lower than about 60% when measured in agastrointestinal simulant, relative to a non-interfering buffer. In someembodiments, the swelling ratio is less than about 2.8, or less thanabout 2.7, or less than about 2.6.

Other diseases that can be treated with the methods, compositions, andkits of the present invention include hypocalcemia, hyperparathyroidism,hungry bone syndrome, depressed renal synthesis of calcitriol, tetanydue to hypocalcemia, renal insufficiency, and ectopic calcification insoft tissues including calcifications in joints, lungs, kidney,conjunctiva, and myocardial tissues Also, the present invention can beused to treat ESRD and dialysis patients, including prophylactictreatment of any of the above.

Also, the polymers described herein can be used as an adjunct to othertherapies e.g. those employing dietary control of phosphorus intake,dialysis inorganic metal salts and/or other polymer resins.

The compositions of the present invention are also useful in removingchloride, bicarbonate, iron ions, oxalate, and bile acids from thegastrointestinal tract. Polymers removing oxalate ions find use in thetreatment of oxalate imbalance disorders, such as such as oxalosis orhyperoxaluria that increases the risk of kidney stone formation.Polymers removing chloride ions find use in treating acidosis,heartburn, acid reflux disease, sour stomach or gastritis, for example.In some embodiments, the compositions of the present invention areuseful for removing fatty acids, bilirubin, and related compounds. Someembodiments may also bind and remove high molecular weight moleculeslike proteins, nucleic acids, vitamins or cell debris.

The present invention provides methods, pharmaceutical compositions, andkits for the treatment of animal. The term “animal” or “animal subject”as used herein includes humans as well as other mammals. One embodimentof the invention is a method of removing phosphate from thegastrointestinal tract of an animal by administering an effective amountof at least one of the crosslinked amine polymers described herein.

The term “treating” and its grammatical equivalents as used hereinincludes achieving a therapeutic benefit and/or a prophylactic benefit.By therapeutic benefit is meant eradication, amelioration, or preventionof the underlying disorder being treated. For example, in ahyperphosphatemia patient, therapeutic benefit includes eradication oramelioration of the underlying hyperphosphatemia. Also, a therapeuticbenefit is achieved with the eradication, amelioration, or prevention ofone or more of the physiological symptoms associated with the underlyingdisorder such that an improvement is observed in the patient,notwithstanding that the patient may still be afflicted with theunderlying disorder. For example, administration of crosslinked aminepolymers, described herein, to a patient suffering from renalinsufficiency and/or hyperphosphatemia provides therapeutic benefit notonly when the patient's serum phosphate level is decreased, but alsowhen an improvement is observed in the patient with respect to otherdisorders that accompany renal failure and/or hyperphosphatemia likeectopic calcification and renal osteodystrophy. For prophylacticbenefit, for example, the crosslinked amine polymers may be administeredto a patient at risk of developing hyperphosphatemia or to a patientreporting one or more of the physiological symptoms ofhyperphosphatemia, even though a diagnosis of hyperphosphatemia may nothave been made. For example, the polymers of the invention may beadministered to a patient with chronic kidney disease wherehyperphosphatemia has not been diagnosed.

The dosages of the polymer, e.g., crosslinked amine polymers, in animalswill depend on the disease being, treated, the route of administration,and the physical characteristics of the animal being treated. In someembodiments in which crosslinked amine polymers are used, the dosagelevels of the crosslinked amine polymers for therapeutic and/orprophylactic uses can be from about 1 gm/day to about 30 gm/day. It ispreferred that these polymers are administered along with meals. Thepolymers may be administered one time a day, two times a day, or threetimes a day. The preferred dosage range is from about 2 gm/day to about20 gm/day and an even preferred dosage range is from about 3 gm/day toabout 7 gm/day. The dose of the polymers described herein can be lessthan about 50 gm/day, preferably less than about 40 gm/day, even morepreferably less than about 30 gm/day, even more preferred less thanabout 20 gm/day, and most preferred is less than about 10 gm/day.

Preferably, the ion-binding polymers, e.g., crosslinked amine polymers,used for therapeutic and/or prophylactic benefits can be administeredalone or in the form of a pharmaceutical composition as describedherein. For example, crosslinked amine polymers of the present inventionmay be co-administered with other active pharmaceutical agents dependingon the condition being treated. Examples of pharmaceutical agents thatmay be co-administered include, but are not limited to, proton pumpinhibitors, calcimimetics (for example, cinacalcet), Vitamin D andanalogs thereof, and phosphate binders. Examples of suitable phosphatebinders include, but are not limited to, aluminum carbonate, calciumcarbonate, calcium acetate (PhosLo), lanthanum carbonate (Fosrenol), andRenagel. This co-administration can include simultaneous administrationof the two agents in the same dosage form, simultaneous administrationin separate dosage forms, and separate administration. For example, forthe treatment of hyperphosphatemia, the crosslinked amine polymers maybe co-administered with calcium salts which are used to treathypocalcemia resulting from hyperphosphatemia. The calcium salt and thepolymer can be formulated together in the same dosage form andadministered simultaneously. Alternatively, the calcium salt and thepolymer can be simultaneously administered, wherein both the agent arepresenting separate formulation. In another alternative, the calciumsalt can be administered just followed by the polymer, or vice versa. Inthe separate administration protocol, the polymer and calcium salt maybe administered a few minutes apart, or a few hours apart, or a few daysapart.

The polymer can be administered by injection, topically, orally,transdermally, or rectally. Preferably, the polymer or thepharmaceutical composition comprising the polymer is administeredorally. The oral form in which the polymer is administered can includepowder, tablet, capsule, solution, or emulsion. The effective amount canbe administered in a single dose or in a series of doses separated byappropriate time intervals, such as hours.

The invention also provides methods of removing anionic pollutants fromwastewater by contacting the wastewater with an anion-binding polymer ofthe invention, where anionic pollutants, e.g., phosphate, are adsorbedto the polymer.

V. Kits

In still another aspect, the present invention provides kits for thetreatment of anion imbalance disorders, e.g., for the treatment ofphosphate imbalance disorders. These kits comprise a polymer or polymersdescribed herein and instructions teaching the use of the kit accordingto the various methods and approaches described herein. Such kits mayalso include information, such as scientific literature references,package insert materials, clinical trial results, and/or summaries ofthese and the like, which indicate or establish the activities and/oradvantages of the composition. Such information may be based on theresults of various studies, for example, studies using experimentalanimals involving in vivo models and studies based on human clinicaltrials. Kits described herein can be provided, marketed and/or promotedto health providers, including physicians, nurses, pharmacists,formulary officials, and the like. Kits for cosmetic use may beprovided, marketed and/or promoted directly to consumers.

All publications and patent applications mentioned in this specificationare herein incorporated by reference to the same extent as if eachindividual publication or patent application was specifically andindividually indicated to be incorporated by reference.

It will be apparent to one of ordinary skill in the art that manychanges and modification can be made to the disclosures presented hereinwithout departing from the spirit or scope of the appended claims.

EXAMPLES Example 1 Phosphate Binding Measurement Protocols

In this Example various protocols for measuring the capacity of apolymer for binding of an anion (in this case, phosphate) are described.

Phosphate Binding Capacity Measurements in a Non Interfering Buffer

An aliquot of dried polymer of weight P(gr), was mixed under gentleagitation with a fixed volume, V(L), of the following buffer: 20 mMH₃PO4, 80 mM NaCl, 100 mM MES sodium salt (morpholinoethanesulfonicacid) and a pH of 6.5. When single binding measurements were made, thelatter buffer was used. When multiple measurements were made, e.g., forthe plotting of a binding isotherm, the phosphate concentration of thebuffer was varied. The starting phosphate ion concentration is referredto as P_(start)(mM). The solution can be referred to as anon-interfering buffer as it contains no other competing solutes thatcompete with the phosphate ions for binding to the polymer resin. Afterresin equilibration, the solution was decanted by centrifugation and thesupernatant analyzed for residual phosphate concentration by ionicchromatography, P_(eq)(mM). The binding capacity was calculated asV*(P_(start)-P_(eq))/P expressed in mmol/gr as indicated in the tablesfor the corresponding polymers.

Binding Capacity in a Gastrointestinal Simulant

This procedure was designed to mimic the conditions of use of aphosphate binding polymer in a GI tract and measure the bindingcharacteristics of the polymer for phosphate (target solute) in thepresence of other metabolites (competing solutes). A liquid meal wasartificially digested in the presence of pepsin and pancreatic juice toproduce gastrointestinal (GI) simulant. The sequence of addition ofenzymes and the pH profile were controlled so that the digestion processwas simulated down to the jejunum level:

The following components were added one at a time in the followingorder: Powder Milk 291 g, Beneprotein 72.8 g, Dextrose 152 g, Polycose156 g, NaCl 17.6 g to ˜2.5 L of ddH₂O until they dissolved (they werestirred vigorously, but foaming was avoided). After the NaCl haddissolved 240 g of Corn Oil was added. Then the volume was brought up to4 L with ddH₂O. The mixture was stirred vigorously for 2 hours. At thistime pH was ˜6.4. Then, 153 ml of 3M HCl was added drop wise to a finalpH of 2.0 (˜150 ml). The mixture was stirred for 15 minutes, after whichtime the pH rose to ˜2.1. Then 800 ml of pepsin in 10 mM HCl was addedto a final concentration of 1 mg/ml. The mixture was stirred at RT for30 minutes, after which time the pH was ˜2.3. Then 5 L of a stocksolution of Pancreatin and Bile Salts in 100 mM NaHCO₃, pH8.4 was addedto a final concentration of 0.3 mg/ml Pancreatin and 2 mg/ml Bile Salts.The mixture was stirred for 120 minutes at room temperature, after whichthe pH was ˜6.5. Meal mimic was stored at −80° C. for up to one monthbefore use.

An aliquot of the GI simulant was centrifuged and the supernatantassayed for phosphate. The phosphate binding assay was like the onedescribed above with non-interfering buffer, except that liquid of theGI simulant was used.

Binding Capacity in ex-vivo Aspirates

Using a tube placed in the lumen of the small intestine, healthypatients were given a meal of the same composition as the one preparedfor the GI simulant described above and aliquots of chyme were thensampled.

Subjects were intubated with a double lumen polyvinyl tube with amercury-weighted bag attached to the end of the tube to facilitatemovement of the tube into the small intestine. Using fluoroscopy todirect placement, one aspiration aperture of the double lumen tube waslocated in the stomach, and the other aperture was at the Ligament ofTreitz (in the upper jejunum).

After correct tube placement, 550 mL of the liquefied test meal(supplemented with a marker, polyethylene glycol (PEG)-2 g/550 mL) wasinfused into the stomach through the gastric aperture at a rate of 22 mLper minute. It required approximately 25 minutes for the entire meal toreach the stomach, simulating the duration of time required to eatnormal meals.

Jejunal chyme was aspirated from the tube whose lumen was located at theLigament of Treitz. This fluid was collected continuously during 30minute intervals for a two and a half hour period. This resulted in 5specimens that were mixed, measured for volume, and lyophilized.

A phosphate binding assay was carried out on the ex-vivo aspirates. Thephosphate binding procedure was like the one described above withnon-interfering buffer, except that the ex-vivo aspirate liquid was used(after reconstitution of the freeze-dried material in the proper amountof de-ionized water). The phosphate binding capacities in the ex-vivoaspirate was calculated in the same way as in GI simulant experiments.

Example 2 Libraries of Crosslinked Polymers Formed in a Bulk SolutionProcess and Measurement for Phosphate Binding Capacity

Creation of Polymer Libraries

The following five examples each comprise a library comprising up to 24crosslinked polymers. Polymers were prepared in batch reactors arrangedin a 4×6 array format. Each reactor had either a 350 microliters or a 3ml volume, was magnetically stirred, and temperature-controlled. In atypical procedure, amine, crosslinkers, solvents and optionally basewere dispensed robotically in each reactor, optionally under agitation.The reactors were then sealed and heated up to the indicated temperaturefor 15 hours. The reactor array was then dismounted and plugs ofcrosslinked polymers transferred in glass vials, ground, washedrepeatedly with de-ionized water, and lyophilized. The five librariesare identified below in Table 3 along with the corresponding reactionconditions used in their creation.

TABLE 3 Library Reaction temper- Reactor volume Example identificationature (° C.) (microliters) 1 100275 85 350 2 100277 60 350 3 100279 80350 4 100353 80 350 5 100384 80 3000

Phosphate Binding Capacity Measurements in a Non Interfering Buffer

Binding capacities for phosphate ion were also determined for each ofthe polymers of the libraries. See Example 1 for procedure.

Results

Tables 4-8 provide materials and the quantities used in forming thepolymers of each of the 5 libraries, along with the measured phosphatebinding capacities in a non-interfering buffer for the polymers formed.Entries correspond to the weight of chemicals used in each reaction wellin mg, along with the phosphate binding capacity of the polymer gelobtained (blank indicates no crosslinked gel was formed in thatparticular reaction).

TABLE 4 Library: Plate3 (ID: 100275) Unit: mg Phosphate B-SM- X- bindingRow Col water 22-DA Cl-3 NaOH DMSO (mmol/gr) 1 1 128.51 67.74 51.63 9.140.00 1 2 130.70 57.94 61.82 10.94 0.00 1 3 132.33 50.61 69.43 12.29 0.001 4 133.59 44.93 75.33 13.33 0.00 3.042 1 5 134.60 40.39 80.04 14.170.00 0 1 6 135.43 36.69 83.89 14.85 0.00 0 2 1 136.42 32.26 88.50 15.660.00 3.703 2 2 137.05 29.41 91.45 16.19 0.00 3.624 2 3 137.58 27.0393.93 16.63 0.00 2.858 2 4 138.03 25.00 96.03 17.00 0.00 2.566 2 5138.42 23.26 97.84 17.32 0.00 2.761 2 6 138.76 21.74 99.42 17.60 0.002.82 3 1 132.04 64.98 49.52 17.53 34.60 3 2 134.77 55.13 58.82 20.8247.26 3 3 136.79 47.87 65.67 23.25 57.22 3 4 138.34 42.30 70.93 25.1165.27 3.087 3 5 139.57 37.90 75.09 26.58 71.91 2.946 3 6 140.56 34.3278.47 27.78 77.48 2.535 4 1 141.75 30.06 82.48 29.20 79.73 2.674 4 2142.50 27.35 85.04 30.11 90.45 3.038 4 3 143.13 25.09 87.18 30.86 97.982.895 4 4 143.66 23.17 88.99 31.50 103.56 2.571 4 5 144.12 21.52 90.5432.05 107.86 2.636 4 6 0.00 0.00 0.00 0.00 0.00 5.374

TABLE 5 Library: Plate1 (ID: 100277) Unit: mg Phosphate B-SM- X- X-binding Row Col water 20-TeA EP-1 EP-4 DMF (mmol/gr) 1 1 123.69 110.7512.95 0.00 1 2 124.02 107.66 16.36 0.00 0.00 1 3 124.33 104.74 19.590.00 0.00 1 4 124.63 101.98 22.65 0.00 0.00 1 5 124.91 99.35 25.55 0.000.00 4.183 1 6 125.17 96.86 28.31 0.00 0.00 4.237 2 1 125.59 92.98 32.610.00 0.00 4.631 2 2 125.89 90.08 35.81 0.00 0.00 4.594 2 3 126.18 87.3738.81 0.00 0.00 4.667 2 4 126.45 84.81 41.64 0.00 0.00 4.586 2 5 126.7182.40 44.31 0.00 0.00 4.535 2 6 126.95 80.12 46.83 0.00 0.00 4.311 3 10.00 181.12 0.00 34.60 0.00 3 2 0.00 159.58 0.00 47.26 104.77 3 3 0.00142.63 0.00 57.22 118.23 3.112 3 4 0.00 128.93 0.00 65.27 128.56 2.991 35 0.00 117.63 0.00 71.91 136.73 2.798 3 6 0.00 108.15 0.00 77.48 143.353.271 4 1 0.00 104.33 0.00 79.73 148.83 3.258 4 2 0.00 86.08 0.00 90.45156.12 3.062 4 3 0.00 73.27 0.00 97.98 160.76 2.176 4 4 0.00 63.77 0.00103.56 164.62 2.228 4 5 0.00 56.46 0.00 107.86 167.88 2.407 4 6 0.000.00 0.00 0.00 170.67 5.224 4 6 0.00 0.00 0.00 0.00 0.00

TABLE 6 Library: Plate3 (ID: 100279) Unit: mg Phosphate binding Row Colwater B-SM-20-TeA X-Cl-3 X-Cl-2 (mmol/gr) 1 1 123.95 108.47 15.49 0.00 12 124.34 104.88 19.47 0.00 1 3 124.70 101.51 23.19 0.00 1 4 125.04 98.3626.68 0.00 1 5 125.36 95.40 29.97 0.00 3.958 1 6 125.66 92.61 33.06 0.004.309 2 1 126.13 88.30 37.82 0.00 4.417 2 2 126.47 85.14 41.33 0.004.424 2 3 126.78 82.19 44.59 0.00 4.392 2 4 127.08 79.44 47.64 0.004.407 2 5 127.36 76.87 50.49 0.00 4.14 2 6 127.62 74.46 53.16 0.00 4.3143 1 0.00 118.41 0.00 26.19 3 2 0.00 102.78 0.00 29.56 3 3 0.00 90.800.00 32.14 3 4 0.00 81.32 0.00 34.18 3 5 0.00 73.64 0.00 35.84 3 6 0.0067.28 0.00 37.21 2.237 4 1 0.00 58.81 0.00 39.03 2.403 4 2 0.00 53.430.00 40.19 2.704 4 3 0.00 48.96 0.00 41.15 2.614 4 4 0.00 45.17 0.0041.97 1.714 4 5 0.00 41.93 0.00 42.67 2.294 4 6 0.00 0.00 0.00 0.005.295

TABLE 7 Library: Plate1 (ID: 100353) Unit: mg Phosphate B-SM- B-SM-binding Row Col 20-TeA 22-DA X-Cl-3 NaOH (mmol/gr) 1 1 142.77 11.1433.97 24.05 1 2 117.71 9.19 44.82 31.73 1 3 100.13 7.82 52.42 37.125.838 1 4 87.12 6.80 58.05 41.10 5.38 1 5 77.10 6.02 62.39 44.17 5.549 16 69.15 5.40 65.83 46.61 5.826 2 1 64.71 5.05 67.75 47.97 5.452 2 257.99 4.53 70.66 50.03 3.358 2 3 52.54 4.10 73.01 51.70 3.45 2 4 48.023.75 74.97 53.08 4.27 2 5 44.22 3.45 76.61 54.24 3.469 2 6 40.98 3.2078.02 55.24 4.058 3 1 111.71 26.16 39.87 28.23 3 2 89.37 20.93 51.0436.14 3 3 74.48 17.44 58.49 41.41 5.154 3 4 63.85 14.95 63.81 45.185.784 3 5 55.87 13.08 67.80 48.01 5.596 3 6 49.66 11.63 70.91 50.205.287 4 1 46.24 10.83 72.62 51.42 5.261 4 2 41.13 9.63 75.17 53.23 4.7434 3 37.04 8.67 77.22 54.67 4.076 4 4 33.69 7.89 78.90 55.86 3.924 4 530.90 7.24 80.29 56.85 2.896 4 6 0.00 0.00 0.00 0.00 5.287

TABLE 8 Library: Plate1 (ID: 100384) Unit: mg Phosphate binding Row ColX-Cl-3 B-SM-22-DA water NaOH (mmol/gr) 1 1 643.88 422.44 1752.36 227.941 2 692.40 378.56 1743.80 245.12 4.362 1 3 731.79 342.94 1736.85 259.064.09 1 4 764.40 313.44 1731.10 270.61 3.198 1 5 791.85 288.62 1726.26280.33 2.951 1 6 815.27 267.44 1722.12 288.62 2.005 2 1 643.88 422.441752.36 227.94 2 2 692.40 378.56 1743.80 245.12 2 3 731.79 342.941736.85 259.06 2 4 764.40 313.44 1731.10 270.61 4.794 2 5 791.85 288.621726.26 280.33 2 6 815.27 267.44 1722.12 288.62 4.332 3 1 643.88 422.441752.36 227.94 3 2 692.40 378.56 1743.80 245.12 3 3 731.79 342.941736.85 259.06 3 4 764.40 313.44 1731.10 270.61 4.511 3 5 791.85 288.621726.26 280.33 5.086 3 6 815.27 267.44 1722.12 288.62 4.61 4 1 643.88422.44 1752.36 227.94 4 2 692.40 378.56 1743.80 245.12 4 3 731.79 342.941736.85 259.06 4 4 764.40 313.44 1731.10 270.61 4 5 791.85 288.621726.26 280.33 4.816 4 6 0.00 0.00 0.00 0.00 5.17

Example 3 Synthesis of 1,3-Diaminopropane/epichlorohydrin CrosslinkedBeads Formed in a Suspension Process

A 3-liter reaction vessel was used, comprising a three necked roundbottom flask with four side baffles. The reaction flask was equippedwith an oil heating bath, cold-water reflux condenser, and mechanicalstirrer with a 3 inch propeller. To this reaction vessel was introduceda solution of 1,3-diaminopropane (90.2 g, 1.21 mole) dissolved in 90.2 gof water, surfactant (branched dodecylbenzene sulfonic acid sodium salt,6.4 g dissolved in 100 g of water) and 1 Kg of toluene. This initialcharge was agitated to 600 rpm for 2 minutes and then lowered to 300 rpmfor 10 minutes before the epichlorohydrin was added. The 300 rpm speedwas maintained through out the remainder of the experiment. The solutionwas heated to 80° C. and also maintained at this temperature through outthe experiment.

In a separate vessel, a 40 mass % solution of epichlorohydrin in toluenewas prepared. Using a syringe pump, 1.2 equivalents of epichlorohydrin(134.7 g, (1.45 mole)) were added to the initial charge reaction vesselover a 3 hour period. The reaction was continued for an additional 2hours before adding 0.75 equivalents of sodium hydroxide (36.5 g (0.91mole)) in a 40 weight % solution. The sodium hydroxide solution wasadded to the reaction via a syringe pump over a 2.5 hour period. Thereaction was maintained at 80° C. for a further 8 hours.

After this time, beads that formed were purified by removing thetoluene, washing with 1000 ml of acetone, followed by methanol, a 20%solution of NaOH (to remove the surfactant), and then twice more withdeionized water. The beads were freeze dried for 3 days to give a finewhite powder weighing at 160 g (92% yield) and having a mean diameter of93 μm.

Example 4 Synthesis of 1,3-Diaminopropane/1,3-DichloropropaneCrosslinked Polymer

Using water as solvent, 1000 mg of B-SM-22-DA was mixed with 1524 mg ofX-Cl-3 and 2524 mg of water in a 20 mL scintillation vial. The reactionwas subjected to magnetic stirring and maintained at a temperature of80° C. overnight, followed by a temperature of 90° C. for two additionalhours. A 34 wt. % of reaction mixture (1716 mg) was purified by 3washing in water/centrifugation steps and gave 144.7 mg of powder of thepolymer of the present example.

Example 5 Synthesis of 1,3-Diaminopropane/1,3-DichloropropaneCrosslinked Polymer

Using water as a solvent, 2000 mg of B-SM-22-DA was mixed with 3048 mgof X-C1'and 5048 mg of water in a 20 mL scintillation vial. The reactionwas subjected to magnetic stirring and maintained at a temperature of80° C. overnight.

3597 mg of NaOH solution at 30 wt. % in water was added after 3 hours ofreaction to scavenge the acid formed during the reaction as thecrosslinker used was an alkylhalide. A 20.3 wt. % of reaction mixture(2773.5 mg) was purified by 3 washing in water/centrifugation steps andgave 591.3 mg of powder of the polymer of the present example.

Example 6 Synthesis of Crosslinked Beads Prepared with1,3-Diaminopropane/1,3-dichloropropane Using a Prepolymer Approach

Preparation of Pre-Polymer

The reaction vessel used was a 250 mL, two necked round bottom flask,equipped with a cold-water reflux condenser, magnetic stirrer, and runover an argon atmosphere. To this reaction vessel is introduced asolution of 1,3-diaminopropane (31.15 g, 0.42 mole) dissolved in 30.15 gof water. This initial charge is agitated to 300 rpm. The solution washeated to 80° C. and maintained at this temperature through out theexperiment. Using a syringe pump, 1 equivalent (47.47 g, 40.0 mL, 0.42mol) of 1,3 dichloropropane (Aldrich 99%) was added over a 2-hourperiod. The reaction was continued for an additional 2 hours beforeadding 10 mole % (with respect to 1,3-diaminopropane) of sodiumhydroxide (1.68 g (0.042 mole) of NaOH and made up to a 40 weight %solution of water). The sodium hydroxide solution was added to thereaction via pipette over a 2 minute period. The reaction was maintainedat 80° C. for a further 4 hours. The solution at 80° C. is viscous andupon cooling to 25° C. becomes a solid plug that is readily soluble inwater.

Purification

To the solid plug water is added, washing with 200 ml of water and 200mL of MeOH. This is then added to a 1 L beaker that contains a 50/50solution of MeOH/Isopropyl alcohol. The white polymer precipitates.After placing the suspension into a centrifuge, the supernatant liquidis removed. This process is repeated using isopropyl alcohol a further 2times. The white precipitate is then dried under reduced pressure atroom temperature to remove the isopropyl alcohol. Weight of polymerisolated: Mn (GPC relative to polyethylenimine standard) ˜600.

Synthesis Crosslinked Particles

The white pre-polymer (8.7 g) was placed into a flask with 1.3 g ofbranched dodecylbenzene sulfonic acid sodium salt (30 wgt % solution inwater) and 34.8 g of toluene. This gave a 20 weight % solution ofpolymer suspended in toluene. The polymer was ground to micron sizedparticles with a mechanical grinder (Brand: IKA. Model: Ultra-Turax T8).2.2 g of the resulting suspension was loaded into a 10 mL reaction flaskequipped with a heater, a mechanical stirrer, and a syringe pump. Thereaction flask was charged with an additional 3779 mg of toluene. Theflask was heated to 80° C. and the stirrer was turned on (500 RPM).After 3 hours of stirring at this temperature, 112.2 mg (0.0012 mole) ofepichlorohydrin was added over a 1.5-hour period. The reaction wasallowed to proceed a further 2 hours before the addition of 224.4 mg(0.0056 mol) of sodium hydroxide (in a 40 weight % solution of water),which was delivered over a 2 hour period. The reaction was allowed tocool to room temperature and the stirring was stopped. The beads werepurified by removing the toluene, washing with methanol, and then a 20%solution of NaOH (to remove the surfactant) and twice more withdeionized water. The beads were freeze dried for 3 days to give a finewhite powder. The binding capacity measured in a non interfering bufferwas 3.85 mmol/gr.

Example 7 Synthesis and Isolation of Low Molecular Weight Polymer(Prepolymer) Prepared with 1,3-Diaminopropane/1,3-dichloropropane 1

Abbreviations used in the following examples:

Epichlorohydrin: ECH

N,N,N′,N′-tetrakis (3-aminopropyl) 1,4 diaminobutane: BTA

BC: binding capacity

In this Example, the effect of varying the ratio of monomer (in thiscase, a prepolymer) to solvent in the reaction mix on binding capacityand swelling ratio was shown. This Example describes a process involvingtwo parts: first, the synthesis of a soluble prepolymer adduct from 1,3diaminopropane and 1,3dichloropropane, and second, the preparation ofinsoluble beads by further crosslinking of the prepolymer by ECH. Thesecond reaction consisted of an inverse suspension process wherein thewater to prepolymer ratio was varied. The impact of this variation onbinding performance and swelling was evaluated.

Synthesis of Prepolymer

Step 1 (Preparation of pre-polymer): The reaction vessel used was a 250mL, two necked round bottom flask, equipped with a cold-water refluxcondenser, magnetic stirrer and run over an argon atmosphere. To thisreaction vessel was introduced a solution of 1,3-diaminopropane (31.15g, 0.42 mole) dissolved in 30.15 g of water. This initial charge wasagitated to 300 rpm. The solution was heated to 80° C. and maintained atthis temperature through out the experiment. Using a syringe pump, 1equivalent (47.47 g, 40.0 mL, 0.42 mol) of 1,3 dichloropropane (Aldrich99%) was added over a 2 hour period. The reaction was continued for anadditional 2 hours before adding 10 mole % (with respect to1,3-diaminopropane) of sodium hydroxide (1.68 g (0.042 mole) of NaOH andmade up to a 40 weight % solution of water). The sodium hydroxidesolution was added to the reaction via pipette over a 2 minute period.The reaction was maintained at 80° C. for a further 4 hours. Thesolution at 80° C. was viscous and upon cooling to 25° C. became a solidplug that was readily soluble in water.

Step 2 (Purification): Water was added to the solid plug, washing with200 ml of water and 200 mL of MeOH. This was then added to a 1 L beakerthat contains a 50/50 solution of MeOH/Isopropyl alcohol. The whitepolymer precipitated. After centrifugation, the supernatant liquid wasremoved. This process was repeated using isopropyl alcohol a further 2times. The white precipitate was then dried under reduced pressure atroom temperature to remove the isopropyl alcohol. Molecular weight ofpolymer isolated: Mn (GPC relative to polyethyleneimine standard) ˜600.

Synthesis of Micron Sized, Crosslinked Particles Prepared with1,3-Diaminopropane/1,3 Dichloropropane Pre-Polymer in a Semi-Continuous24 Well, Parallel Polymerization Reactor.

The white pre-polymer 1 (8.7 g) was placed into a flask with 1.3 g ofbranched dodecylbenzene sulfonic acid sodium salt (30 wt % solution inwater) and 34.8 g of toluene. This gave a 20 weight % solution ofpolymer suspended in toluene. The emulsion was ground to micron sizeddroplets with a high shear homogenizer (Brand: IKA. Model: Ultra-TuraxT8). 2.2 g of the resulting emulsion was loaded into 24 of the 10 mLreaction flasks of the reactor which was equipped with a heater, amechanical stirrer and a syringe pump. Into each reaction flask wascharged an additional 3779 mg of toluene. The flasks were heated to 80°C. and the stirrer was turned on (500 RPM). Water was loaded into thetubes in an amount necessary to produce various ratios of prepolymer towater. After 3 hours of stirring at this temperature, the desired amountof epichlorohydrin (in this Example, epichlorohydrin was added to anamount equal to 20% of the dry weight of the pre-polymer) was added overa 1.5 hour period. The reaction was allowed to proceed a further 2 hoursbefore the addition of 224.4 mg, (0.0056 mol) of sodium hydroxide (in a40 weight % solution of water), which was delivered over a 2 hourperiod. The reaction was allowed to cool to room temperature and thestirring was stopped. The beads were purified by removing the toluene,washing with methanol and then a 20% solution of NaOH (to remove thesurfactant) and then with HCl to protonate the bead. The beads were thenwashed twice with deionized water to remove excess HCl. The beads werefreeze dried for 3 days to give a fine white powder.

The polymer beads thus synthesized were analyzed for binding capacity(BC) in non-interfering buffer and in a GI simulant, and for swellingratio. The results are summarized in Table 9.

TABLE 9 1,3 diamino propane/1,3 dichloropropane/ECH gel beads. Effect ofthe monomer to water ratio on Binding Capacity and Swelling Swelling BCBC (g of Monomer (mmol/gr) (mmol · gr) H20/g to water Non GI of ratioInterfering simulant polymer) 1.67 3.85 1.54 2.92 1.42 3.68 1.43 3.341.25 3.61 1.34 3.50 1.11 3.55 1.34 3.70 0.83 3.31 1.16 5.22 0.55 2.900.91 14.00

These results show that the binding capacities in both non-interferingbuffer and in GI simulant increase as the monomer to water ratioincreases, while the swelling ratio decreases and reaches to the desiredrange.

Example 8 Synthesis of Micron Sized, Crosslinked Particles from GroundBTA/ECH Bulk Gel Using a 24 Well Parallel Polymerization Reactor

In this Example the effect of varying the amount of crosslinker relativeto monomer on binding capacity and swelling ratio was shown.

The following stock solution was prepared: 2 molar equivalents ofconcentrated HCl was added to 1 molar equivalent of BTA over a 2 hourperiod. Water was then added to the solution such that the resultingsolution achieved the following weight % composition: BTA 45 weight %HCl 10 weight %, water 45 weight %. Into each flask of a 24 well reactorusing 5 mL flasks was placed 0.6 g of the prepared stock solution. Thedesired amount of epichlorohydrin to achieve the monomer:crosslinkerratio to be tested was added to each vial. The reactor was heated to 80°C. for 9 hours. The reactor was allowed to cool. To each vial was addedwater to swell the resulting gel. The gel was then ground to micronsized particles with a high shear homogenizer (Brand: IKA. Model:Ultra-Turax T8). The particles were purified by removing the water,washing with methanol and then a 20% solution of NaOH and then with HClto protonate the amine functionalized particle. The particles were thenwashed twice with deionized water to remove excess HCl. The particleswere freeze dried for 3 days to give a fine white powder.

The results of binding capacity and swelling studies are summarized inTable 10.

TABLE 10 BTA/ECH gel: Data on Swelling and Binding Capacities againstcrosslinker content. Bulk Gels (Monomer to water ratio is 75 wt %Bow-Tie (2HCl) in water). Monomer to water ratios ranges from 3.5(ECH:BTA = 0.85) to 4.8 (ECH:BTA = 6.4) Swelling BC BC ratio (gr.ECH:BTA (mmol/gr) (mmol · gr) Water/ Mole Non GI gr. ratio interferingsimulant Polymer) 0.70 0.00 0.00 0.85 2.23 0.35 1.00 2.46 0.49 16.681.15 2.57 0.49 10.98 1.30 2.84 0.58 6.15 1.45 2.91 0.65 4.69 1.60 2.910.77 3.85 1.79 2.88 0.85 3.13 1.98 0.00 0.98 2.77 2.00 2.46 1.00 2.552.00 2.46 1.00 2.55 2.16 2.73 0.99 2.46 2.35 2.67 0.96 2.20 2.40 2.170.93 1.97 2.40 2.17 0.93 1.97 2.80 1.86 0.82 1.81 2.80 1.86 0.82 1.813.20 1.63 0.73 1.84 3.20 1.63 0.73 1.84 3.60 1.28 0.64 1.57 3.60 1.280.64 1.57 4.00 1.09 0.58 1.57 4.00 1.09 0.58 1.57 4.40 0.88 0.45 2.034.40 0.88 0.45 2.03 4.90 0.42 0.35 1.47 4.90 0.42 0.35 1.47 5.40 0.420.28 1.50 5.40 0.42 0.28 1.50 5.90 0.07 0.27 1.55 5.90 0.07 0.27 1.556.40 0.06 0.22 1.55 6.40 0.06 0.22 1.55

These data show that the binding capacity in the GI simulant goesthrough a maximum as the crosslinker to amine ratio is varied. In thisparticular system the optimum binding capacity in the GI stimulant isobserved at a crosslinker ratio of 1.8 to 2.8, corresponding to a NCvalue of 3.6 to 5.6 respectively. Within that range of crosslinking theswelling ratio is minimal. Similar tests routinely may be carried outfor other monomers and crosslinkers using this polymerization protocolto determine the ratio that gives the desired results for the particularuse to which the polymer will be put.

Example 9 Synthesis of Micron Sized, Crosslinked BTA/ECH Beads viaInverse Suspension

The following stock solution was prepared: 2 molar equivalents ofconcentrated HCl was added to 1 molar equivalent of BTA over a 2 hourperiod. Water and surfactant (branched dodecylbenzene sulfonic acidsodium salt, 30 weight % in water) was then added to the solution suchthat the resulting solution achieved the following weight % composition:BTA 41.8 weight %, HCl 9.4 weight %, water 41.1 weight %, surfactant (30weight % in water) 7.7 weight %.

The reaction vessel used was a 0.25 liter, three necked round bottomflask with four side baffles, equipped with an oil heating bath,cold-water reflux condenser and mechanical stirrer with a 1 inchpropeller. To this reaction vessel was introduced 25 g of the preparedstock solution and 75 g of toluene.

Into a separate vessel, a 40 mass % solution of epichlorohydrin intoluene was prepared. Using a syringe pump, the desired amount of ECHwas added over a 90 minute period. The reaction was continued for anadditional 2 hours before beginning a dehydration using a dean starkapparatus. The reaction end point was reached when all the water fromthe system had been removed. The beads were purified by removing thetoluene, washing with methanol and then a 20% solution of NaOH (toremove the surfactant) and then with HCl to protonate the bead. Thebeads were then washed twice with deionized water to remove excess HCl.The beads were freeze dried for 3 days to give a fine white powder.

The results of binding capacity and swelling studies are summarized inTable 11.

TABLE 11 BTA/ECH gel beads: Swelling and Binding Capacities againstcrosslinker content Swelling ECH: BC BC ratio (gr. BTA (mmol/gr)(mmol/gr) Water/ Mole Non Digested gr. ratio interfering meal Polymer)1.00 2.50 0.58 25.29 1.00 2.77 0.55 13.01 1.25 2.97 0.65 7.69 1.25 3.030.61 7.07 1.50 3.13 0.71 4.41 1.50 3.14 0.69 3.99 1.75 3.13 0.78 3.061.75 3.10 0.87 3.41 2.00 3.07 0.99 3.13 2.00 2.80 1.00 2.82 2.00 2.820.73 3.17 2.50 2.76 1.03 2.48 3.00 2.56 0.82 2.40 3.50 0.00 0.71 2.283.00 2.32 0.70 2.25 3.00 2.61 0.80 2.03 3.50 2.81 0.59 1.85 4.00 0.000.58 1.99 4.00 2.19 0.77 1.93 4.50 2.11 0.30 1.99 5.00 1.96 0.55 1.72

These results show that the binding capacity in the GI simulant goesthrough a maximum as the crosslinker to amine molar ratio is varied. Inthis particular system the optimum binding capacity in the GI stimulantis observed at a crosslinker ratio of 1.75 to 3, corresponding to a NCvalue of 3.5 to 6 respectively Within that range of eros slinking theswelling ratio is minimal. Similar tests routinely may be carried outfor other monomers and crosslinkers using this polymerization protocolto determine the ratio that gives the desired results for the particularuse to which the polymer will be put.

Example 10 Synthesis of Micron Sized, Crosslinked Particles from GroundPolyallylamine/ECH Bulk Gel Using a 24 Well Parallel PolymerizationReactor

This Example illustrates the synthesis of a polymer using a highmolecular weight monomer and varying monomer to water ratios in thereaction mixture. The conditions employed were identical to thosedescribed in Example 8, except that polyallylamine (Mw=60,000 g/mole)was used instead of BTA. The ECH to allylamine repeat unit ratio was1:0.106 (corresponding to a NC of 2.2). The initial polyallyamine towater ratio was varied from 1:1 to 1.4. As a comparative example,crosslinked polyallylamine isolated from Renagel tablets was used.

Swelling Amine BC BC ratio (gr. to water (mmol/gr) (mmol/gr) Water/ MoleNon Digested gr. ratio interfering meal Polymer) 0.20 3.66 0.92 19.000.33 4.12 1.36 6.00 0.50 4.20 1.62 4.00 Renagel 3.85 1.40 9.00

These data indicate that a higher amine to water ratio led to a smallerswelling ratio and was accompanied by a higher binding capacity in theGI simulant. Similar tests routinely may be carried out for othermonomers and crosslinkers using this polymerization protocol todetermine the ratio that gives the desired results for the particularuse to which the polymer will be put.

Example 11 Measure of the Binding Interference Level

This example illustrates the nieasuiement of binding inteiference, usinga polymer of the invention and, for comparison, a prior art polymer. Acrosslinked polyamine material (EC172A) was prepared according toprotocol described in Example 9, with a ECH:BTA mole ratio of 2.5, and a(BTA+ECH) to water ratio of 1.73 by weight. The binding interference wascompared with Renagel.

The “degree of interference in binding” or “binding interference,” asused herein, refers to the fractional decrease in binding capacity forthe target ion observed between a binding experiment in a noninterfering buffer, and in a gastrointestinal (GI) simulant, at the sameconcentration of target anion in equilibrium. A binding isotherm in anon interfering buffer was first obtained by plotting the bindingcapacity versus the phosphate concentration at equilibrium for a varietyof phosphate concentrations. That isotherm was then fitted by anexponential function to predict the binding capacity at any phosphateconcentration. The binding capacity measured in the GI simulant was thenreported on the same isotherm, plotting the point of phosphateconcentration versus phosphate binding at equilibrium for the GIsimulant and extending a vertical line through this point to intersectthe non-interfering isotherm. The interference degree was then computedas the (BCNI−BCGI)/BCNI*100.

The binding interference is shown for EC127A is shown in the Table belowand in FIG. 3.

Predicted Pstart Peq BC BC Interference (mM) (mM) (mmol/gr) (mmol/gr)(%) 6.25 3.31 1.18 2.17 45.7 6.25 3.28 1.19 2.16 45.0 6.25 3.24 1.212.15 44.0

The binding interference for RENAGEL is shown in the Table below and inFIG. 4.

Predicted Pstart Peq BC BC Interference (mM) (mM) (mmol/gr) (mmol/gr)(%) 6.25 2.70 1.42 4.53 68.7 6.25 2.54 1.48 4.46 66.7

The binding interference foi EC172A is about 34% lower than that ofRENAGEL.

Example 12 Ion Binding Properties in Human Ex-Vivo Aspirates

A crosslinked polyamine material (EC172A) was prepared according toprotocol described in Example 9, with a ECH:BTA mole ratio of 2.5, and a(BTA+ECH) to water ratio of 1.73 by weight. The material was then testedfor phosphate binding in a human aspirate collected as described inExample 1.

The binding of phosphate of EC172A was compared to that of crosslinkedpolyallyamine active pharmaceutical isolated from Renagel (Genzyme).EC172A exhibits a much lower level of interference, as well as a muchlower index of swelling (2.5 vs. 9 for Renagel)

Avg Avg BC SD Predicted % Peq SD (mmol/ (mmol/ BC inter- (mM) (mM) gr)gr) (mmol/gr) ference Renagel API 2.37 0.01 1.32 0.00 4.37 70 EC172A1.55 0.04 1.64 0.02 1.68 2.5

In another experiment both materials, EC172A and Renagel, were used in adifferent human ex-vivo aspirate to quantitate the degree ofinterference on phosphate binding produced by competing solutes such ascitrate anions and bile acids. Citrate anions and bile acids weretitrated by ion chromatography and enzymatic assay respectively. Datashown below (mean of six volunteers) indicate that the polymer of thepresent invention exhibit much better selectivity and overall binding ofphosphate.

BC BC (Bile BC [PO4] (PO4) [citrate] (citrate) Acid) (Bile) mM mmol/g mMmmol/g mM mmol/g Control 5.722 1.667 4.928 (no polymer) Renagel 3.0191.078 0.596 0.429 1.32 1.443 EC172A 1.78 1.573 1.316 0.141 4.65 0.109

Example 13 Gel Porosity Measurement Using the Solute PartitioningTechnique

This Example illustrates measurement of gel porosity. The measurementswere carried out on a polymer of the invention, and on a commerciallyavailable phosphate-binding polymer for comparison As a polymer of theinvention, a crosslinked polyamine material (EC172A) was preparedaccording to protocol described in Example 9, with a ECH:BTA mole ratioof 2.5, and a (BTA+ECH) to water ratio of 1.73 by weight. Forcomparison, the same porosity measurements were carried out on Renagel.

The probes were 8 polyethyleneglycols (PEG) of MW ranging from 200 to20,000 Da and 4 polyethyleneoxides (PEO) (30,000 to 230,000 Da).

All probes were dissolved in 30 mM ammonium acetate buffer pH 5.5(concentration 5 g/L). The probe solutions were added to preweighedEC172A HCl wash (5 mL/g) and Renagel HCl wash (15 mL/g dry gel); thenshaken for 4 days on a Vortexer.

Probe solutions were diluted 10× before LC analysis using Polymer LabEvaporative Light Scattering Detector (in order to be in linear range ofdetector guaranteeing that peak area ratio is equal to weightconcentration ratio).

Calculation of Non-accessiblevolume=m_(sw)+[1−c_(before)/c_(after)]m_(solv); where

-   -   m_(sw) water amount uptaken by gel [g/g dry gel]    -   m_(solv) water amount, in which the probe was dissolved at the        beginning [g/g dry gel]    -   C_(before) and c_(after):concentrations of probe before and        after equilibrium. The ratio c_(before)/c_(after) is equal to        ratio of the peak area obtained with LC analysis.

The results of this comparative Example are shown in FIGS. 5 and 6; FIG.5 illustiates the results in terms of molecular weight while FIG. 6illustrates the results in terms of size of the solutes. EC172A showsconstant molecular exclusion for solutes down to a MW of 200, ascompared with Renagel, which demonstrates decreasing exclusion at MW ashigh as 1000.

Example 14 Post Modification of Beads with Chloropropylamine,Hydrochloride

Preparation of Stock Solution:

-   -   Chloropropylamine, hydrochloride (B-SM-34-A) in water at 50 wt.        %−d=1.132    -   Sodium hydroxide in water at 30 wt. % (by dilution of a 50 wt. %        solution)−d=1.335

Synthesis:

FR-0005-144, a phosphate binder polymei prepared according to Example 9,with a ECH:BTA mole ratio of 2.5, and a (BTA+ECH) water ratio of 1.73 byweight, was used as a substrate fox further aminification: TheFR-0005-144 beads were transferred to 4 mL-vials (two 4×6 platescontaining each 21 vial) and water, chioropropylamine, hydrochloridestock solution and sodium hydroxide stock solution were added using aliquid dispensing robot. Vials were sealed with a cap, and plates wereset up on reactors equipped with a heating system and individualStirling

Heating and stirring were turned on for 12 hours: Reactor's temperaturewas set at 85° C. and stirring rate at 1200 rpm.

Purification:

Each material was transferred to disposable culture tubes (16×100 mm)and washed once with methanol, twice with an hydrochloric acid solutionin water at 1 M, and three times with water. Beads were separated eachtime by centrifugation.

They were then dried in a lyophilizer and analyzed for Digested Mealscreen, Non-Interfering buffer and swelling ratio. The results are shownbelow in Table 12 and in FIG. 7.

TABLE 12 Characteristics of polymers prepared by post modification ofbeads with chloropropylamine, hydrochloride Swelling ratio (g B-SM-34-ANaOH mol. of B-SM-34- wt. ratio vs Ratio (vs. B- BC DM screen BC NIScreen water/g FR-0005-144 water A NaOH FR-0005-144 SM-34-DA) (mmol/gr)(mmol/g) of gel) 222.1 864.5 22.2 1.71 0.1 0.25 0.94 2.84 2.91 233.3883.0 46.7 3.59 0.2 0.25 0.91 2.94 2.69 203.7 749.0 61.1 4.70 0.3 0.250.95 2.85 2.83 209.1 746.3 83.6 6.43 0.4 0.25 0.97 2.91 2.64 209 723.5104.5 8.04 0.5 0.25 0.97 2.89 2.58 0 0.0 0.0 0.00 227 761.3 136.2 10.480.6 0.25 0.96 2.90 2.60 235 762.8 164.5 12.65 0.7 0.25 1.00 2.97 2.67231.3 725.9 185.0 14.23 0.8 0.25 0.99 2.88 2.86 278.5 844.1 250.7 19.280.9 0.25 0.99 2.90 3.38 236.2 690.4 236.2 18.17 1.0 0.25 1.00 2.96 2.730 0.0 0.0 0.00 204.1 792.9 20.4 3.14 0.1 0.5 0.92 2.81 2.85 271 1021.554.2 8.34 0.2 0.5 0.95 2.81 2.74 247 902.5 74.1 11.40 0.3 0.5 0.97 2.852.85 225.5 797.9 90.2 13.87 0.4 0.5 0.97 2.93 2.61 238.2 815.4 119.118.32 0.5 0.5 1.01 2.84 2.68 270.7 0.0 0.0 0.00 0.89 2.73 2.98 199.7660.5 119.8 18.43 0.6 0.5 0.98 2.91 2.70 230.6 736.1 161.4 24.83 0.7 0.51.01 3.03 2.46 221.3 680.9 177.0 27.23 0.8 0.5 0.98 2.92 2.58 212.5629.3 191.3 29.42 0.9 0.5 1.02 3.04 2.61 200.4 570.4 200.4 30.83 1.0 0.51.06 2.93 2.46 0 0 0 0 213.1 826.17 21.3 4.92 0.1 0.75 0.94 2.80 2.92203.7 764.66 40.7 9.40 0.2 0.75 0.94 2.81 2.82 212.4 771.18 63.7 14.700.3 0.75 0.97 2.84 3.04 218.2 765.38 87.3 20.14 0.4 0.75 1.00 2.88 2.99203.4 688.43 101.7 23.47 0.5 0.75 1.03 2.90 2.64 0 0 0.0 0.00 214.3698.95 128.6 29.67 0.6 0.75 1.05 2.94 2.50 228.8 718.09 160.2 36.95 0.70.75 1.04 2.95 2.60 235.2 709.23 188.2 43.41 0.8 0.75 1.08 3.02 2.55216.8 627.06 195.1 45.02 0.9 0.75 1.00 2.95 2.65 206.7 572.41 206.747.69 1.0 0.75 1.00 3.03 2.48 0 0 0.0 0.00 199.7 772.69 20.0 6.14 0.11.0 0.97 2.75 2.85 206.4 771.62 41.3 12.70 0.2 1.0 0.97 2.77 3.30 216779.26 64.8 19.94 0.3 1.0 0.98 2.83 2.93 213.3 741.63 85.3 26.25 0.4 1.01.00 2.85 3.43 212.9 712.4 106.5 32.75 0.5 1.0 1.04 2.95 2.66 193.3 00.0 0.00 0.95 2.73 2.95 240.6 773.63 144.4 44.41 0.6 1.0 1.02 2.94 2.88294.5 908.43 206.2 63.42 0.7 1.0 1.07 2.94 2.58 214.1 632.43 171.3 52.690.8 1.0 1.06 3.05 2.60 205.5 580.15 185.0 56.90 0.9 1.0 1.08 3.04 2.66201.2 541.7 201.20 61.90 1.0 1.0 1.09 3.07 2.91 0 0 0.00 0.00

Example 15 Synthesis of Phosphate-Templated Micron Sized, CrosslinkedParticles from N,N′(tetra-3-aminopropyl) 1,4diaminobutane/epichlorohydrin

The following stock solution was prepared: 1 molar equivalent ofPhosphoric acid (Aldrich, 85 wgt % in water) was added to 1 molarequivalent of N,N′ (tetra-3-aminopropyl) 1,4 diaminobutane over a 2 hourperiod. Water was then added to the solution such that the resultingsolution achieved the following weight % composition: N,N′(tetra-3-aminopropyl) 1,4 diaminobutane 42 weight %, H₃PO₄ 13 weight %,water 45 weight %. The reactor contained 24 wells used 5 mL flasks, eachflask containing a magnetic stir bar. Into each flask was placed 0.6-0.7g of the prepared stock solution. The stirrers were on. The desiredamount of epichlorohydrin was added neat to each vial. The reactor washeated to 60° C. for 1 hour and then heated to 80° C. for 8 hours. Thereactor was allowed to cool. To each vial was added water to swell theresulting gel. The gel was transferred to a 4×6 plate with 10 mL testtubes. The gel was then ground to micron sized particles with amechanical grinder (Brand: IKA. Model: Ultra-Turax T8). The particleswere purified by removing the water, washing with methanol and furtherwashing with a 20% solution of NaOH. The gel particles were subsequentlywashed with 1.0 molar HCl, mixed for 30 minutes, then the gel wasallowed to settle and the supernatant liquid was decanted off. Thisprocess was repeated 5 times to protonate the amine functionalizedparticle with Chloride and replace the bound H₃PO₄. The gel particleswere then washed with a 20% solution of NaOH to deprotonate the aminefunctionalized gel particles. The gel particles were then washed twicewith deionized water to remove excess NaOH/NaCl. The gel particles werefreeze dried for 3 days to give a fine white powder. The synthesis issummarized in Table 13.

TABLE 13 Synthesis of gels that have been Molecular imprinted withPhosphoric acid. ID 102776 B- SM- X-EP- 20- B-SM- X- 1/B- Gel TeA 20-TeAphosphoric water EP-1 X-EP-1 B-SM-20- SM-20- Present Row Col (mg)(Moles) acid (mg) (mg) (mg) (Moles) TeA/H3PO4 TeA in well 1.0 1.0 347.50.0011 107.7 369.8 71.1 0.0008 1.00 0.70 x 1.0 2.0 339.5 0.0011 105.2361.4 79.4 0.0009 1.00 0.80 x 1.0 3.0 337.7 0.0011 104.6 359.4 88.80.0010 1.00 0.90 ✓ 1.0 4.0 352.1 0.0011 109.1 374.8 102.9 0.0011 1.001.00 ✓ 1.0 5.0 355.4 0.0011 110.1 378.2 114.3 0.0012 1.00 1.10 ✓ 1.0 6.0366.1 0.0012 113.4 389.6 128.4 0.0014 1.00 1.20 ✓ 2.0 1.0 355.3 0.0011110.1 378.1 135.0 0.0015 1.00 1.30 ✓ 2.0 2.0 338.6 0.0011 104.9 360.4138.6 0.0015 1.00 1.40 ✓ 2.0 3.0 356.2 0.0011 110.4 379.1 156.2 0.00171.00 1.50 ✓ 2.0 4.0 349.7 0.0011 108.3 372.2 163.5 0.0018 0.99 1.61 ✓2.0 5.0 342.2 0.0011 106.0 364.2 170.0 0.0018 1.00 1.70 ✓ 2.0 6.0 351.40.0011 108.9 374.1 184.9 0.0020 1.00 1.80 ✓ 3.0 1.0 364.1 0.0012 112.8387.5 212.8 0.0023 1.00 2.00 ✓ 3.0 2.0 351.2 0.0011 108.8 373.8 246.40.0027 1.00 2.40 ✓ 3.0 3.0 358.3 0.0011 111.0 381.4 293.2 0.0032 1.002.81 ✓ 3.0 4.0 340.2 0.0011 105.4 362.1 318.2 0.0034 1.00 3.20 ✓ 3.0 5.0368.9 0.0012 114.3 392.6 388.2 0.0042 1.00 3.59 ✓ 3.0 6.0 360.5 0.0011111.7 383.7 421.5 0.0046 1.00 4.00 ✓ 4.0 1.0 345.3 0.0011 107.0 367.5444.0 0.0048 1.00 4.40 ✓ 4.0 2.0 364.0 0.0012 112.8 387.4 510.7 0.00551.00 4.80 ✓ 4.0 3.0 351.2 0.0011 108.8 373.7 533.7 0.0058 1.00 5.20 ✓4.0 4.0 365.5 0.0012 113.2 389.0 598.3 0.0065 0.99 5.63 ✓ 4.0 5.0 358.50.0011 111.1 381.6 628.8 0.0068 1.00 6.02 ✓

The polymers synthesized as described above bind phosphate.

While preferred embodiments of the present invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the invention. It should be understoodthat various alternatives to the embodiments of the invention describedherein may be employed in practicing the invention. It is intended thatthe following claims define the scope of the invention and that methodsand structures within the scope of these claims and their equivalents becovered thereby.

1. A pharmaceutical composition comprising a particulate crosslinkedamine polymer as an active ingredient and a pharmaceutically acceptableexcipient, the particulate crosslinked amine polymer particles having achloride content of less than about 20 mol % of the amine group contentand a size in the range of 5 to 500 microns, the crosslinked aminepolymer comprising repeat units derived from polymerization of a non-polymeric amine and a crosslinking agent such that the polymer has aswelling ratio of less than about 5 as measured in a physiologicalmedium, the non-polymeric amine being selected from the group consistingof (i) a non-polymeric amine of formula (I)

wherein each n, independently, is equal to or greater than 3; m is equalto or greater than 1; and each R_(1,) independently, is H or optionallysubstituted alkyl or aryl or is linked to a neighboring R₁ to form anoptionally substituted alicyclic, aromatic, or heterocyclic group;provided the non-polymeric amine of formula I is not 1,3-diaminopropane,and (ii) a non-polymeric amine of formula (IV)

wherein each n, independently, is equal to or greater than 3; each r,independently, is 0, 1, or 2; and each R₁, independently, is H oroptionally substituted alkyl or aryl or is linked to a neighboring R₁ toform an optionally substituted alicyclic, aromatic, or heterocyclicgroup.
 2. The pharmaceutical composition of claim 1 wherein thenon-polymeric amine is a non-polymeric amine of formula IV.
 3. Thepharmaceutical composition of claim 2 wherein the non- polymeric amineis

and n is 3, 4 or
 5. 4. The pharmaceutical composition of claim 3 whereinn is
 3. 5. The pharmaceutical composition of claim 3 wherein n is
 5. 6.The pharmaceutical composition of claim 3 wherein the polymer particleshave a size in the range of 25 to 250 microns.
 7. The pharmaceuticalcomposition of 3 wherein the polymer particles are spherical beadshaving a diameter of 25 to 250 microns.
 8. The pharmaceuticalcomposition of claim 1 wherein the polymer particles have a size in therange of 25 to 250 microns.
 9. The pharmaceutical composition of 1wherein the polymer particles are spherical beads having a diameter of25 to 250 microns.
 10. The pharmaceutical composition of claim 1 whereinthe crosslinking agent is 1,3-dichloropropane or epichlorohydrin. 11.The pharmaceutical composition of claim 1 wherein the crosslinking agentis epichlorohydrin.
 12. The pharmaceutical composition of claim 2wherein the polymer particles have a size in the range of 25 to 250microns.
 13. The pharmaceutical composition of 2 wherein the particlesare spherical beads having a diameter of 25 to 250 microns.
 14. Thepharmaceutical composition of claim 2 wherein the crosslinking agent is1,3-dichloropropane or epichlorohydrin.
 15. The pharmaceuticalcomposition of claim 2 wherein the crosslinking agent isepichlorohydrin.
 16. The pharmaceutical composition of claim 1 whereinthe polymer has a gel pore volume distribution where less than about 20%of the pore volume of the gel is accessible to non-interacting solutesof molecular weight greater than about 200 as measured in aphysiological medium.
 17. The pharmaceutical composition of claim 3wherein the polymer has a gel pore volume distribution where less thanabout 20% of the pore volume of the gel is accessible to non-interactingsolutes of molecular weight greater than about 200 as measured in aphysiological medium.
 18. The pharmaceutical composition of claim 1wherein the polymer binds phosphate ion in vivo with a binding capacityof greater than 0.5 mmol/g.
 19. The pharmaceutical composition of claim3 wherein the polymer binds phosphate ion in vivo with a bindingcapacity of greater than 0.5 mmol/g.
 20. The pharmaceutical compositionof claim 1 wherein the polymer has a gel pore volume distribution whereless than about 20% of the pore volume of the gel is accessible tonon-interacting solutes of molecular weight greater than about 200 asmeasured in a physiological medium, and the polymer binds phosphate ionin vivo with a binding capacity of greater than 0.5 mmol/g.
 21. Thepharmaceutical composition of claim 3 wherein the polymer has a gel porevolume distribution where less than about 20% of the pore volume of thegel is accessible to non-interacting solutes of molecular weight greaterthan about 200 as measured in a physiological medium, and the polymerbinds phosphate ion in vivo with a binding capacity of greater than 0.5mmol/g.
 22. The pharmaceutical composition of claim 1 wherein thepolymer particles have a chloride content of less than about 5 mol % ofthe amine group content.
 23. The pharmaceutical composition of claim 1wherein the polymer particles are substantially chloride-free.
 24. Thepharmaceutical composition of claim 1 wherein the composition is in theform of a powder.
 25. The pharmaceutical composition of claim 3 whereinthe composition is in the form of a powder.
 26. The pharmaceuticalcomposition of claim 1 wherein the composition is contained within asachet.
 27. The pharmaceutical composition of claim 3 wherein thecomposition is contained within a sachet.
 28. The pharmaceuticalcomposition of claim 1 wherein the composition is in the form of asterile packaged powder.
 29. The pharmaceutical composition of claim 3wherein the composition is in the form of a sterile packaged powder. 30.The pharmaceutical composition of claim 1 wherein the composition is inthe form of a soft or hard gelatin capsule.
 31. The pharmaceuticalcomposition of claim 3 wherein the composition is in the form of a softor hard gelatin capsule.
 32. A pharmaceutical composition comprising aparticulate crosslinked amine polymer as an active ingredient and apharmaceutically acceptable excipient, the particulate crosslinked aminepolymer particles being spherical beads having a size in the range of 5to 500 microns, the crosslinked amine polymer comprising repeat unitsderived from polymerization of a non-polymeric amine and a crosslinkingagent such that the polymer has a swelling ratio of less than about 5 asmeasured in a physiological medium, the non-polymeric amine beingselected from the group consisting of (i) a non-polymeric amine offormula (I)

wherein each n, independently, is equal to or greater than 3; m is equalto or greater than 1; and each R₁, independently, is H or optionallysubstituted alkyl or aryl or is linked to a neighboring R₁ to form anoptionally substituted alicyclic, aromatic, or heterocyclic group;provided the non-polymeric amine of formula I is not 1,3-diaminopropane,and (ii) a non-polymeric amine of formula (IV)

wherein each n, independently, is equal to or greater than 3; each r,independently, is 0, 1, or 2; and each R_(1,) independently, is H oroptionally substituted alkyl or aryl or is linked to a neighboring R₁ toform an optionally substituted alicyclic, aromatic, or heterocyclicgroup.
 33. The pharmaceutical composition of claim 32 wherein the non-polymeric amine is a non-polymeric amine of formula IV.
 34. Thepharmaceutical composition of claim 33 wherein the non- polymeric amineis

and n is 3, 4 or
 5. 35. The pharmaceutical composition of claim 34wherein n is
 3. 36. The pharmaceutical composition of claim 34 wherein nis
 5. 37. The pharmaceutical composition of claim 34 wherein the polymerparticles have a size in the range of 25 to 250 microns.
 38. Thepharmaceutical composition of claim 33 wherein the crosslinking agent is1,3-dichloropropane or epichlorohydrin.
 39. The pharmaceuticalcomposition of claim 33 wherein the crosslinking agent isepichlorohydrin.
 40. The pharmaceutical composition of claim 32 whereinthe polymer has a gel pore volume distribution where less than about 20%of the pore volume of the gel is accessible to non-interacting solutesof molecular weight greater than about 200 as measured in aphysiological medium.
 41. The pharmaceutical composition of claim 34wherein the polymer has a gel pore volume distribution where less thanabout 20% of the pore volume of the gel is accessible to non-interactingsolutes of molecular weight greater than about 200 as measured in aphysiological medium.
 42. The pharmaceutical composition of claim 32wherein the composition is in the form of a powder.
 43. Thepharmaceutical composition of claim 34 wherein the composition is in theform of a powder.
 44. The pharmaceutical composition of claim 34 whereinthe composition is contained within a sachet.
 45. The pharmaceuticalcomposition of claim 34 wherein the composition is contained within asachet.
 46. The pharmaceutical composition of claim 32 wherein thecomposition is in the form of a sterile packaged powder.
 47. Thepharmaceutical composition of claim 34 wherein the composition is in theform of a sterile packaged powder.
 48. The pharmaceutical composition ofclaim 32 wherein the composition is in the form of a soft or hardgelatin capsule.
 49. The pharmaceutical composition of claim 34 whereinthe composition is in the form of a soft or hard gelatin capsule.